The secreted immune factor, Tumor necrosis factor alpha (TNF-α), is an important inflammatory mediator that helps neutralize and destroy infectious agents as well as cancer cells. In fact, it is the second-most studied gene in the human genome, second to the tumor suppressor p53 (1). However, as the saying goes, “Too much of a good thing is bad for you.” Indeed, chronic high-level production of TNF-α is associated with autoimmune diseases such as plaque psoriasis, rheumatoid arthritis, and inflammatory bowel disease (e.g., Crohn’s disease and ulcerative colitis).
This persistent high level of TNFα leads to damaging and life-threatening effects against normal tissue, such as the resultant sepsis due to bowel perforations in inflammatory bowel disease or irreversible and disabling joint damage seen in rheumatoid arthritis. Importantly, antibody-based drugs that neutralize TNF-α have dramatically altered the symptomology of sufferers of autoimmune diseases, leading to significant improvements in their quality of life.
The ominous name, TNF-α, suggests that it is a major factor that can cause the necrotic death cancer cells. However, in most cases, TNF-α is used by the cancer cell as a survival factor involving the induction of gene networks activated by transcription factors belonging to the NF-κB family. This may explain in part how chronic viral and bacterial infections (and the associated TNF-α production) can lead to cancer formation. Recombinant TNF-α had long ago been touted as the magic bullet to kill cancer cells, but the toxicity of systemically administered TNF-α towards normal tissue and its inability to broadly kill cancer cells led to the demise of this approach.
Due to developments over the past couple decades, the contrasting duality of action of TNF-α is now finally explained by the presence or absence of key enzymes from the inhibitor of apoptosis (IAP) family, particularly the two redundant proteins cellular IAP1 (cIAP1 or BIRC2 gene) and cellular IAP2 (cIAP2 or BIRC3 gene). The cIAPs modify key TNF-α signaling effectors with ubiquitin chains with branching structures that either form protein scaffolds to recruit other signaling molecules or mark proteins for degradation by the cell’s garbage can, the proteasome. Recently, class of experimental anti-cancer drugs that bind to cIAP1 and cIAP2, called Smac mimetics (SMs), have been developed that trigger the ubiquitination and degradation of those two cIAPs, thus shifting TNF-α signaling fate in SM-treated cancer cells from survival to death. These experimental drugs continue to be evaluated in clinical trials.
To further understand how SMs work, we undertook an unbiased screening approach in which we individually targeted over 18,000 genes in breast cancer cells with pools of synthetic RNA duplexes (siRNA) to neutralize each of those specific gene products. We then treated these cells with an SM to cause cell death. The goal of this screen was to identify genes that are necessary for SMs to kill highly aggressive triple-negative breast cancers, which do not respond to conventional anti-estrogen or anti-Her2/neu therapies. This screen identified transcription factor called SP3 (for specificity protein 3), which we found to be required for the cell-autonomous and SM-induced production of TNF-α(2). Without SP3 or TNF-α, the triple-negative breast cancer cells, as well as glioblastoma cells survived SM treatment. The ability of the transcription factor SP3 to enhance the production of TNF-α was shown to be based on the ability of SP3 to bind to the gene promoter of TNF-α and to cooperate with the binding of nearby NF-κB transcription factors. In addition, we observed higher SP3 expression in cancer cells versus normal cells, which may explain why cancer cells are more sensitive to SM-induced killing. Furthermore, a nine-gene signature profile for the TNF-α-inducing transcription factors clearly discriminated cancer from normal cells and thus may act as a potential biomarker of SM action.
Image courtesy of Shawn Beug, Eric LaCasse and Robert Korneluk (Ottawa, Canada).
Our study has demonstrated that the SP3 transcription factor, in concert with NF-κB, has a clear and critical role in producing TNF-α that is needed for SMs to kill many different cancer cell types. The importance of our work extends beyond cancer biology and therapeutics, as any insight into the production of this important inflammatory mediator could lead to better ways to control its production and deleterious effects, especially in cases of chronic TNF-α activation associated with diseases. Alternatively, transient activation of SP3 and TNF-α by novel small molecules could be used for cancer and infection control, especially in combination with an SM.
Hence, SMs have resurrected the concept that TNF-α can be strongly cytotoxic in cancer cells when the cIAPs are pharmacologically targeted. Recent reports, for example, have shown that currently approved cancer immunotherapies (such as immune checkpoint inhibitor biologics and CAR-T engineered adoptive cell therapy) can enhance local TNF-α production by T cells to synergize with SMs and kill cancer cells (3-5).
These findings are described in the article entitled The transcription factor SP3 drives TNF-α expression in response to Smac mimetics, recently published in the journal Science Signaling.
Bibliography
1. Dolgin, E. (2017) The most popular genes in the human genome. Nature 551: 427-431.
2. Beug ST, Cheung HH, Sanda T, St-Jean M, Beauregard CE, Mamady H, Baird SD, LaCasse EC, Korneluk RG. (2019) The transcription factor SP3 drives TNF-α expression in response to Smac mimetics. Science Signaling 12. pii: eaat9563. doi: 10.1126/scisignal.aat9563.
3. Beug, ST, Beauregard CE, Healy C, Sanda T, St-Jean M, Chabot J, Walker DE, Mohan A, Earl N, Lun X, Senger DL, Robbins SM, Staeheli P, Forsyth PA, Alain T, LaCasse EC, Korneluk RG. (2017) Smac mimetics synergize with immune checkpoint inhibitors to promote tumour immunity against glioblastoma. Nature Communications 8. doi: 10.1038/ncomms14278.
4. Kearney CJ, Lalaoui N, Freeman AJ, Ramsbottom KM, Silke J, Oliaro J. (2017) PD-L1 and IAPs co-operate to protect tumors from cytotoxic lymphocyte-derived TNF. Cell Death & Differentiation 24: 1705-1716.
5. Michie J, Beavis PA, Freeman AJ, Vervoort SJ, Ramsbottom KM, Narasimhan V, Lelliott EJ, Lalaoui N, Ramsay RG, Johnstone RW, Silke J, Darcy PK, Voskoboinik I, Kearney CJ, Oliaro J. (2019) Antagonism of IAPs Enhances CAR T-cell Efficacy. Cancer Immunology Research 7: 183-192.