Mutant TRP53 exerts a target gene-selective dominant-negative effect to drive tumor development.

Mutations in Trp53, prevalent in human cancer, are reported to drive tumorigenesis through dominant-negative effects (DNEs) over wild-type TRP53 function as well as neomorphic gain-of-function (GOF) activity. We show that five TRP53 mutants do not accelerate lymphomagenesis on a TRP53-deficient background but strongly synergize with c-MYC overexpression in a manner that distinguishes the hot spot Trp53 mutations. RNA sequencing revealed that the mutant TRP53 DNE does not globally repress wild-type TRP53 function but disproportionately impacts a subset of wild-type TRP53 target genes. Accordingly, TRP53 mutant proteins impair pathways for DNA repair, proliferation, and metabolism in premalignant cells. This reveals that, in our studies of lymphomagenesis, mutant TRP53 drives tumorigenesis primarily through the DNE, which modulates wild-type TRP53 function in a manner advantageous for neoplastic transformation.

BCL-2 protein family: attractive targets for cancer therapy.

Acquired resistance to cell death is a hallmark of cancer. The BCL-2 protein family members play important roles in controlling apoptotic cell death. Abnormal over-expression of pro-survival BCL-2 family members or abnormal reduction of pro-apoptotic BCL-2 family proteins, both resulting in the inhibition of apoptosis, are frequently detected in diverse malignancies. The critical role of the pro-survival and pro-apoptotic BCL-2 family proteins in the regulation of apoptosis makes them attractive targets for the development of agents for the treatment of cancer. This review describes the roles of the various pro-survival and pro-apoptotic members of the BCL-2 protein family in normal development and organismal function and how defects in the control of apoptosis promote the development and therapy resistance of cancer. Finally, we discuss the development of inhibitors of pro-survival BCL-2 proteins, termed BH3-mimetic drugs, as novel agents for cancer therapy.

Impaired ribosome biogenesis checkpoint activation induces p53-dependent MCL-1 degradation and MYC-driven lymphoma death.

MYC-driven B-cell lymphomas are addicted to increased levels of ribosome biogenesis (RiBi), offering the potential for therapeutic intervention. However, it is unclear whether inhibition of RiBi suppresses lymphomagenesis by decreasing translational capacity and/or by p53 activation mediated by the impaired RiBi checkpoint (IRBC). Here we generated Eµ-Myc lymphoma cells expressing inducible short hairpin RNAs to either ribosomal protein L7a (RPL7a) or RPL11, the latter an essential component of the IRBC. The loss of either protein reduced RiBi, protein synthesis, and cell proliferation to similar extents. However, only RPL7a depletion induced p53-mediated apoptosis through the selective proteasomal degradation of antiapoptotic MCL-1, indicating the critical role of the IRBC in this mechanism. Strikingly, low concentrations of the US Food and Drug Administration-approved anticancer RNA polymerase I inhibitor Actinomycin D (ActD) dramatically prolonged the survival of mice harboring Trp53+/+;Eµ-Myc but not Trp53-/-;Eµ-Myc lymphomas, which provides a rationale for treating MYC-driven B-cell lymphomas with ActD. Importantly, the molecular effects of ActD on Eµ-Myc cells were recapitulated in human B-cell lymphoma cell lines, highlighting the potential for ActD as a therapeutic avenue for p53 wild-type lymphoma.

Intact TP-53 function is essential for sustaining durable responses to BH3-mimetic drugs in leukemias.

Selective targeting of BCL-2 with the BH3-mimetic venetoclax has been a transformative treatment for patients with various leukemias. TP-53 controls apoptosis upstream of where BCL-2 and its prosurvival relatives, such as MCL-1, act. Therefore, targeting these prosurvival proteins could trigger apoptosis across diverse blood cancers, irrespective of TP53 mutation status. Indeed, targeting BCL-2 has produced clinically relevant responses in blood cancers with aberrant TP-53. However, in our study, TP53-mutated or -deficient myeloid and lymphoid leukemias outcompeted isogenic controls with intact TP-53, unless sufficient concentrations of BH3-mimetics targeting BCL-2 or MCL-1 were applied. Strikingly, tumor cells with TP-53 dysfunction escaped and thrived over time if inhibition of BCL-2 or MCL-1 was sublethal, in part because of an increased threshold for BAX/BAK activation in these cells. Our study revealed the key role of TP-53 in shaping long-term responses to BH3-mimetic drugs and reconciled the disparate pattern of initial clinical response to venetoclax, followed by subsequent treatment failure among patients with TP53-mutant chronic lymphocytic leukemia or acute myeloid leukemia. In contrast to BH3-mimetics targeting just BCL-2 or MCL-1 at doses that are individually sublethal, a combined BH3-mimetic approach targeting both prosurvival proteins enhanced lethality and durably suppressed the leukemia burden, regardless of TP53 mutation status. Our findings highlight the importance of using sufficiently lethal treatment strategies to maximize outcomes of patients with TP53-mutant disease. In addition, our findings caution against use of sublethal BH3-mimetic drug regimens that may enhance the risk of disease progression driven by emergent TP53-mutant clones.

Strip and Cka negatively regulate JNK signalling during Drosophila spermatogenesis.

One fundamental property of a stem cell niche is the exchange of molecular signals between its component cells. Niche models, such as the Drosophila melanogaster testis, have been instrumental in identifying and studying the conserved genetic factors that contribute to niche molecular signalling. Here, we identify jam packed (jam), an allele of Striatin interacting protein (Strip), which is a core member of the highly conserved Striatin-interacting phosphatase and kinase (STRIPAK) complex. In the developing Drosophila testis, Strip cell-autonomously regulates the differentiation and morphology of the somatic lineage, and non-cell-autonomously regulates the proliferation and differentiation of the germline lineage. Mechanistically, Strip acts in the somatic lineage with its STRIPAK partner, Connector of kinase to AP-1 (Cka), where they negatively regulate the Jun N-terminal kinase (JNK) signalling pathway. Our study reveals a novel role for Strip/Cka in JNK pathway regulation during spermatogenesis within the developing Drosophila testis.

Last but not least: BFL-1 as an emerging target for anti-cancer therapies.

BFL-1 is an understudied pro-survival BCL-2 protein. The expression of BFL-1 is reported in many cancers, but it is yet to be clarified whether high transcript expression also always correlates with a pro-survival function. However, recent applications of BH3-mimetics for the treatment of blood cancers identified BFL-1 as a potential resistance factor in this type of cancer. Hence, understanding the role of BFL-1 in human cancers and how its up-regulation leads to therapy resistance has become an area of great clinical relevance. In addition, deletion of the murine homologue of BFL-1, called A1, in mice showed only minimal impacts on the well-being of these animals, suggesting drugs targeting BFL-1 would exhibit limited on-target toxicities. BFL-1 therefore represents a good clinical cancer target. Currently, no effective BFL-1 inhibitors exist, which is likely due to the underappreciation of BFL-1 as a potential target in the clinic and lack of understanding of the BFL-1 protein. In this review, the roles of BFL-1 in the development of different types of cancers and drug resistant mechanisms are discussed and some recent advances in the generation of BFL-1 inhibitors highlighted.

Putting the STING back into BH3-mimetic drugs for TP53-mutant blood cancers.

TP53-mutant blood cancers remain a clinical challenge. BH3-mimetic drugs inhibit BCL-2 pro-survival proteins, inducing cancer cell apoptosis. Despite acting downstream of p53, functional p53 is required for maximal cancer cell killing by BH3-mimetics through an unknown mechanism. Here, we report p53 is activated following BH3-mimetic induced mitochondrial outer membrane permeabilization, leading to BH3-only protein induction and thereby potentiating the pro-apoptotic signal. TP53-deficient lymphomas lack this feedforward loop, providing opportunities for survival and disease relapse after BH3-mimetic treatment. The therapeutic barrier imposed by defects in TP53 can be overcome by direct activation of the cGAS/STING pathway, which promotes apoptosis of blood cancer cells through p53-independent BH3-only protein upregulation. Combining clinically relevant STING agonists with BH3-mimetic drugs efficiently kills TRP53/TP53-mutant mouse B lymphoma, human NK/T lymphoma, and acute myeloid leukemia cells. This represents a promising therapy regime that can be fast-tracked to tackle TP53-mutant blood cancers in the clinic.

Genome-wide CRISPR screening identifies a role for ARRDC3 in TRP53-mediated responses.

Whole-genome screens using CRISPR technologies are powerful tools to identify novel tumour suppressors as well as factors that impact responses of malignant cells to anti-cancer agents. Applying this methodology to lymphoma cells, we conducted a genome-wide screen to identify novel inhibitors of tumour expansion that are induced by the tumour suppressor TRP53. We discovered that the absence of Arrestin domain containing 3 (ARRDC3) increases the survival and long-term competitiveness of MYC-driven lymphoma cells when treated with anti-cancer agents that activate TRP53. Deleting Arrdc3 in mice caused perinatal lethality due to various developmental abnormalities, including cardiac defects. Notably, the absence of ARRDC3 markedly accelerated MYC-driven lymphoma development. Thus, ARRDC3 is a new mediator of TRP53-mediated suppression of tumour expansion, and this discovery may open new avenues to harness this process for cancer therapy.

Loss-of-Function but Not Gain-of-Function Properties of Mutant TP53 Are Critical for the Proliferation, Survival, and Metastasis of a Broad Range of Cancer Cells.

Mutations in the tumor suppressor TP53 cause cancer and impart poor chemotherapeutic responses, reportedly through loss-of-function, dominant-negative effects and gain-of-function (GOF) activities. The relative contributions of these attributes is unknown. We found that removal of 12 different TP53 mutants with reported GOFs by CRISPR/Cas9 did not impact proliferation and response to chemotherapeutics of 15 human cancer cell lines and colon cancer-derived organoids in culture. Moreover, removal of mutant TP53/TRP53 did not impair growth or metastasis of human cancers in immune-deficient mice or growth of murine cancers in immune-competent mice. DepMap mining revealed that removal of 158 different TP53 mutants had no impact on the growth of 391 human cancer cell lines. In contrast, CRISPR-mediated restoration of wild-type TP53 extinguished the growth of human cancer cells in vitro. These findings demonstrate that LOF but not GOF effects of mutant TP53/TRP53 are critical to sustain expansion of many tumor types. SIGNIFICANCE: This study provides evidence that removal of mutant TP53, thereby deleting its reported GOF activities, does not impact the survival, proliferation, metastasis, or chemotherapy responses of cancer cells. Thus, approaches that abrogate expression of mutant TP53 or target its reported GOF activities are unlikely to exert therapeutic impact in cancer. See related commentary by Lane, p. 211 . This article is featured in Selected Articles from This Issue, p. 201.

BCL-W makes only minor contributions to MYC-driven lymphoma development.

The BH3-mimetic drug Venetoclax, a specific inhibitor of anti-apoptotic BCL-2, has had clinical success for the treatment of chronic lymphocytic leukaemia and acute myeloid leukaemia. Attention has now shifted towards related pro-survival BCL-2 family members, hypothesising that new BH3-mimetic drugs targeting these proteins may emulate the success of Venetoclax. BH3-mimetics targeting pro-survival MCL-1 or BCL-XL have entered clinical trials, but managing on-target toxicities is challenging. While increasing evidence suggests BFL-1/A1 is a resistance factor for diverse chemotherapeutic agents and BH3-mimetic drugs in haematological malignancies, few studies have explored the role of BCL-W in the development, expansion, and therapeutic responses of cancer. Previously, we found that BCL-W was not required for the ongoing survival and growth of various established human Burkitt lymphoma and diffuse large B cell lymphoma cell lines. However, questions remained about whether BCL-W impacts lymphoma development. Here, we show that BCL-W appears dispensable for MYC-driven lymphomagenesis, and such tumours arising in the absence of BCL-W show no compensatory changes to BCL-2 family member expression, nor altered sensitivity to BH3-mimetic drugs. These results demonstrate that BCL-W does not play a major role in the development of MYC-driven lymphoma or the responses of these tumours to anti-cancer agents.