Medicine Oncology

Cancer-related Molecular Pathways

Description

This cluster of papers focuses on the p53 signaling network, exploring its role in cell cycle regulation, tumor suppression, cancer therapy, apoptosis, and the impact of mutations. The papers cover various aspects of p53-MDM2 interaction, cyclin-dependent kinases, and the intricate mechanisms of p53 regulation in the context of cancer research.

Keywords

p53; cell cycle; MDM2; tumor suppression; cancer therapy; apoptosis; CDKs; mutation; regulation; oncogene

The inhibition of replicative DNA synthesis that follows DNA damage may be critical for avoiding genetic lesions that could contribute to cellular transformation. Exposure of ML-1 myeloblastic leukemia cells to … The inhibition of replicative DNA synthesis that follows DNA damage may be critical for avoiding genetic lesions that could contribute to cellular transformation. Exposure of ML-1 myeloblastic leukemia cells to nonlethal doses of the DNA damaging agents, gamma-irradiation or actinomycin D, causes a transient inhibition of replicative DNA synthesis via both G1 and G2 arrests. Levels of p53 protein in ML-1 cells and in proliferating normal bone marrow myeloid progenitor cells increase and decrease in temporal association with the G1 arrest. In contrast, the S-phase arrest of ML-1 cells caused by exposure to the anti-metabolite, cytosine arabinoside, which does not directly damage DNA, is not associated with a significant change in p53 protein levels. Caffeine treatment blocks both the G1 arrest and the induction of p53 protein after gamma-irradiation, thus suggesting that blocking the induction of p53 protein may contribute to the previously observed effects of caffeine on cell cycle changes after DNA damage. Unlike ML-1 cells and normal bone marrow myeloid progenitor cells, hematopoietic cells that either lack p53 gene expression or overexpress a mutant form of the p53 gene do not exhibit a G1 arrest after gamma-irradiation; however, the G2 arrest is unaffected by the status of the p53 gene. These results suggest a role for the wild-type p53 protein in the inhibition of DNA synthesis that follows DNA damage and thus suggest a new mechanism for how the loss of wild-type p53 might contribute to tumorigenesis.
Cells prepare for S phase during the G 1 phase of the cell cycle. Cell biological methods have provided knowledge of cycle kinetics and of substages of G 1 that … Cells prepare for S phase during the G 1 phase of the cell cycle. Cell biological methods have provided knowledge of cycle kinetics and of substages of G 1 that are determined by extracellular signals. Through the use of biochemical and molecular biological techniques to study effects of growth factors, oncogenes, and inhibitors, intracellular events during G 1 that lead to DNA synthesis are rapidly being discovered. Many cells in vivo are in a quiescent state (G 0 ), with unduplicated DNA. Cells can be activated to reenter the cycle during G 1 . Similarly, cells in culture can be shifted between G 0 and G 1 . These switches in and out of G 1 are the main determinants of post-embryonic cell proliferation rate and are defectively controlled in cancer cells.
A putative tumor suppressor locus on the short arm of human chromosome 9 has been localized to a region of less than 40 kilobases by means of homozygous deletions in … A putative tumor suppressor locus on the short arm of human chromosome 9 has been localized to a region of less than 40 kilobases by means of homozygous deletions in melanoma cell lines. This region contained a gene, Multiple Tumor Suppressor 1 ( MTS1 ), that encodes a previously identified inhibitor (p16) of cyclin-dependent kinase 4. MTS1 was homozygously deleted at high frequency in cell lines derived from tumors of lung, breast, brain, bone, skin, bladder, kidney, ovary, and lymphocyte. Melanoma cell lines that carried at least one copy of MTS1 frequently carried nonsense, missense, or frameshift mutations in the gene. These findings suggest that MTS1 mutations are involved in tumor formation in a wide range of tissues.
Familial cancer syndromes have helped to define the role of tumor suppressor genes in the development of cancer. The dominantly inherited Li-Fraumeni syndrome (LFS) is of particular interest because of … Familial cancer syndromes have helped to define the role of tumor suppressor genes in the development of cancer. The dominantly inherited Li-Fraumeni syndrome (LFS) is of particular interest because of the diversity of childhood and adult tumors that occur in affected individuals. The rarity and high mortality of LFS precluded formal linkage analysis. The alternative approach was to select the most plausible candidate gene. The tumor suppressor gene, p53, was studied because of previous indications that this gene is inactivated in the sporadic (nonfamilial) forms of most cancers that are associated with LFS. Germ line p53 mutations have been detected in all five LFS families analyzed. These mutations do not produce amounts of mutant p53 protein expected to exert a trans-dominant loss of function effect on wild-type p53 protein. The frequency of germ line p53 mutations can now be examined in additional families with LFS, and in other cancer patients and families with clinical features that might be attributed to the mutation.
C J Sherr and J M Roberts Howard Hughes Medical Institute, Department of Tumor Cell Biology, St. Jude Children's Research Hospital, Memphis, Tennessee 38104, USA. C J Sherr and J M Roberts Howard Hughes Medical Institute, Department of Tumor Cell Biology, St. Jude Children's Research Hospital, Memphis, Tennessee 38104, USA.
The ability of p53 to activate transcription from specific sequences suggests that genes induced by p53 may mediate its biological role as a tumor suppressor. Using a subtractive hybridization approach, … The ability of p53 to activate transcription from specific sequences suggests that genes induced by p53 may mediate its biological role as a tumor suppressor. Using a subtractive hybridization approach, we identified a gene, named WAF1, whose induction was associated with wild-type but not mutant p53 gene expression in a human brain tumor cell line. The WAF1 gene was localized to chromosome 6p21.2, and its sequence, structure, and activation by p53 was conserved in rodents. Introduction of WAF1 cDNA suppressed the growth of human brain, lung, and colon tumor cells in culture. Using a yeast enhancer trap, a p53-binding site was identified 2.4 kb upstream of WAF1 coding sequences. The WAF1 promoter, including this p53-binding site, conferred p53-dependent inducibility upon a heterologous reporter gene. These studies define a gene whose expression is directly induced by p53 and that could be an important mediator of p53-dependent tumor growth suppression.
In recent years, members of the protein kinase family have been discovered at an accelerated pace. Most were first described, not through the traditional biochemical approach of protein purification and … In recent years, members of the protein kinase family have been discovered at an accelerated pace. Most were first described, not through the traditional biochemical approach of protein purification and enzyme assay, but as putative protein kinase amino acid sequences deduced from the nucleotide sequences of molecularly cloned genes or complementary DNAs. Phylogenetic mapping of the conserved protein kinase catalytic domains can serve as a useful first step in the functional characterization of these newly identified family members.
Uncontrolled cell proliferation is the hallmark of cancer, and tumor cells have typically acquired damage to genes that directly regulate their cell cycles. Genetic alterations affecting p16(INK4a) and cyclin D1, … Uncontrolled cell proliferation is the hallmark of cancer, and tumor cells have typically acquired damage to genes that directly regulate their cell cycles. Genetic alterations affecting p16(INK4a) and cyclin D1, proteins that govern phosphorylation of the retinoblastoma protein (RB) and control exit from the G1 phase of the cell cycle, are so frequent in human cancers that inactivation of this pathway may well be necessary for tumor development. Like the tumor suppressor protein p53, components of this "RB pathway," although not essential for the cell cycle per se, may participate in checkpoint functions that regulate homeostatic tissue renewal throughout life.
Mutations in the evolutionarily conserved codons of the p53 tumor suppressor gene are common in diverse types of human cancer. The p53 mutational spectrum differs among cancers of the colon, … Mutations in the evolutionarily conserved codons of the p53 tumor suppressor gene are common in diverse types of human cancer. The p53 mutational spectrum differs among cancers of the colon, lung, esophagus, breast, liver, brain, reticuloendothelial tissues, and hemopoietic tissues. Analysis of these mutations can provide clues to the etiology of these diverse tumors and to the function of specific regions of p53. Transitions predominate in colon, brain, and lymphoid malignancies, whereas G:C to T:A transversions are the most frequent substitutions observed in cancers of the lung and liver. Mutations at A:T base pairs are seen more frequently in esophageal carcinomas than in other solid tumors. Most transitions in colorectal carcinomas, brain tumors, leukemias, and lymphomas are at CpG dinucleotide mutational hot spots. G to T transversions in lung, breast, and esophageal carcinomas are dispersed among numerous codons. In liver tumors in persons from geographic areas in which both aflatoxin B 1 and hepatitis B virus are cancer risk factors, most mutations are at one nucleotide pair of codon 249. These differences may reflect the etiological contributions of both exogenous and endogenous factors to human carcinogenesis.
After DNA damage, many cells appear to enter a sustained arrest in the G 2 phase of the cell cycle. It is shown here that this arrest could be sustained … After DNA damage, many cells appear to enter a sustained arrest in the G 2 phase of the cell cycle. It is shown here that this arrest could be sustained only when p53 was present in the cell and capable of transcriptionally activating the cyclin-dependent kinase inhibitor p21. After disruption of either the p53 or the p21 gene, γ radiated cells progressed into mitosis and exhibited a G 2 DNA content only because of a failure of cytokinesis. Thus, p53 and p21 appear to be essential for maintaining the G 2 checkpoint in human cells.
Multiple genetic changes occur during the evolution of normal cells into cancer cells. This evolution is facilitated in cancer cells by loss of fidelity in the processes that replicate, repair, … Multiple genetic changes occur during the evolution of normal cells into cancer cells. This evolution is facilitated in cancer cells by loss of fidelity in the processes that replicate, repair, and segregate the genome. Recent advances in our understanding of the cell cycle reveal how fidelity is normally achieved by the coordinated activity of cyclin-dependent kinases, checkpoint controls, and repair pathways and how this fidelity can be abrogated by specific genetic changes. These insights suggest molecular mechanisms for cellular transformation and may help to identify potential targets for improved cancer therapies.
Mutations in the p53 tumor suppressor are the most frequently observed genetic alterations in human cancer. The majority of the mutations occur in the core domain which contains the sequence-specific … Mutations in the p53 tumor suppressor are the most frequently observed genetic alterations in human cancer. The majority of the mutations occur in the core domain which contains the sequence-specific DNA binding activity of the p53 protein (residues 102-292), and they result in loss of DNA binding. The crystal structure of a complex containing the core domain of human p53 and a DNA binding site has been determined at 2.2 angstroms resolution and refined to a crystallographic R factor of 20.5 percent. The core domain structure consists of a beta sandwich that serves as a scaffold for two large loops and a loop-sheet-helix motif. The two loops, which are held together in part by a tetrahedrally coordinated zinc atom, and the loop-sheet-helix motif form the DNA binding surface of p53. Residues from the loop-sheet-helix motif interact in the major groove of the DNA, while an arginine from one of the two large loops interacts in the minor groove. The loops and the loop-sheet-helix motif consist of the conserved regions of the core domain and contain the majority of the p53 mutations identified in tumors. The structure supports the hypothesis that DNA binding is critical for the biological activity of p53, and provides a framework for understanding how mutations inactivate it.
MDM2 binds the p53 tumor suppressor protein with high affinity and negatively modulates its transcriptional activity and stability. Overexpression of MDM2, found in many human tumors, effectively impairs p53 function. … MDM2 binds the p53 tumor suppressor protein with high affinity and negatively modulates its transcriptional activity and stability. Overexpression of MDM2, found in many human tumors, effectively impairs p53 function. Inhibition of MDM2-p53 interaction can stabilize p53 and may offer a novel strategy for cancer therapy. Here, we identify potent and selective small-molecule antagonists of MDM2 and confirm their mode of action through the crystal structures of complexes. These compounds bind MDM2 in the p53-binding pocket and activate the p53 pathway in cancer cells, leading to cell cycle arrest, apoptosis, and growth inhibition of human tumor xenografts in nude mice.
Mitogen-dependent progression through the first gap phase (G1) and initiation of DNA synthesis (S phase) during the mammalian cell division cycle are cooperatively regulated by several classes of cyclin-dependent kinases … Mitogen-dependent progression through the first gap phase (G1) and initiation of DNA synthesis (S phase) during the mammalian cell division cycle are cooperatively regulated by several classes of cyclin-dependent kinases (CDKs) whose activities are in turn constrained by CDK inhibitors (CKIs). CKIs that govern these events have been assigned to one of two families based on their structures and CDK targets. The first class includes the INK4 proteins (inhibitors of CDK4), so named for their ability to specifically inhibit the catalytic subunits of CDK4 and CDK6. Four such proteins [p16 (Serrano et al. 1993), p15 (Hannon and Beach 1994), p18 (Guan et al. 1994; Hirai et al. 1995), and p19 (Chan et al. 1995; Hirai et al. 1995)] are composed of multiple ankyrin repeats and bind only to CDK4 and CDK6 but not to other CDKs or to D-type cyclins. The INK4 proteins can be contrasted with more broadly acting inhibitors of the Cip/Kip family whose actions affect the activities of cyclin D-, E-, and A-dependent kinases. The latter class includes p21 (Gu et al. 1993; Harper et al. 1993; El-Deiry et al. 1993; Xiong et al. 1993a; Dulic et al. 1994; Noda et al. 1994), p27 (Polyak et al. 1994a,b; Toyoshima and Hunter 1994), and p57 (Lee et al. 1995; Matsuoka et al. 1995), all of which contain characteristic motifs within their amino-terminal moieties that enable them to bind both to cyclin and CDK subunits (Chen et al. 1995, 1996; Nakanishi et al. 1995; Warbrick et al. 1995; Lin et al. 1996; Russo et al. 1996). Based largely on in vitro experiments and in vivo overexpression studies, CKIs of the Cip/Kip family were initially thought to interfere with the activities of cyclin D-, E-, and A-dependent kinases. More recent work has altered this view and revealed that although the Cip/Kip proteins are potent inhibitors of cyclin Eand A-dependent CDK2, they act as positive regulators of cyclin Ddependent kinases. This challenges previous assumptions about how the G1/S transition of the mammalian cell cycle is governed, helps explain some enigmatic features of cell cycle control that also involve the functions of the retinoblastoma protein (Rb) and the INK4 proteins, and changes our thinking about how either p16 loss or overexpression of cyclin D-dependent kinases contribute to cancer. Here we focus on the biochemical interactions that occur between CKIs and cyclin Dand E-dependent kinases in cultured mammalian cells, emphasizing the manner by which different positive and negative regulators of the cell division cycle cooperate to govern the G1-to-S transition. To gain a more comprehensive understanding of the biology of CDK inhibitors, readers are encouraged to refer to a rapidly emerging but already extensive literature (for review, see Elledge and Harper 1994; Sherr and Roberts 1995; Chellappan et al. 1998; Hengst and Reed 1998a; Kiyokawa and Koff 1998; Nakayama 1998; Ruas and Peters 1998).
STRUCTURED ABSTRACT Importance Li-Fraumeni Syndrome (LFS) is an heterogenous cancer predisposition caused by pathogenic TP53 variants, characterized by a lifelong high risk of a broad spectrum of cancers. At least … STRUCTURED ABSTRACT Importance Li-Fraumeni Syndrome (LFS) is an heterogenous cancer predisposition caused by pathogenic TP53 variants, characterized by a lifelong high risk of a broad spectrum of cancers. At least certain pathogenic TP53 variants have been shown be immunogenic in a somatic context. Whether neoantigenicity contributes to the heterogeneity of LFS is unknown. Objective To analyze the correlations between predicted neoantigenic properties of pathogenic TP53 missense variants and LFS patterns. Design Association study between predicted MHC-I presentation scores for TP53 hotspot variants, LFS presentation and individual HLA-I genotyping in carriers of TP53 germline pathogenic variants using data from mutation databases and clinical registries. Setting MHC-I presentation scores were generated for the set of nonameric neo-peptides surrounding each TP53 missense variant against 145 different HLA-I using NetMHCpan 4.1 and the Allele Frequency Net Database. Genotype-phenotype data were leveraged from the public TP53 database (germline dataset, n=3,446; https://tp53.isb-cgc.org/ ) and two independent LFS clinical registries (n=339). Individual correlations between HLA-I genotyping, TP53 missense variants and phenotypes were investigated in a group of 173 subjects with LFS. Participants Individuals with LFS enrolled in LFS-registries in France and Germany were included. Main outcomes and measures A predicted neoantigenic score (PNS) was calculated for each variant. Correlations with median age at first cancer, and cancer type were analyzed. Results Among 709 carriers of frequent TP53 pathogenic variants, PNS was strongly correlated with median age at first cancer (range 18-43 years, p=0.01319, R=0.69). Compared to carriers of low PNS (<1) variants, carriers of high PNS (>2) variants showed delayed median age at first diagnosis (34 [CI95%, 29-40] vs. 25 years [CI95%, 22-27]), fewer sarcomas (osteosarcoma [RR 0.29, p-adj = 0.02]; soft-tissue [RR 0.41, p-adj = 0.02]), and more cancer types typically not associated with LFS [RR 1.61, p-adj = 0.02]. Conclusion and Relevance MHC-I neoantigenic properties of TP53 variants modify cancer risk and spectrum in carriers of pathogenic TP53 variants, suggesting that individual variant-specific immune response may contribute to the heterogenous presentation of LFS.
Cyclin F, a noncanonical member of the cyclin protein family, plays a critical role in regulating transitions in the cell division cycle. Unlike canonical cyclins, which bind and activate cyclin-dependent … Cyclin F, a noncanonical member of the cyclin protein family, plays a critical role in regulating transitions in the cell division cycle. Unlike canonical cyclins, which bind and activate cyclin-dependent kinases (CDKs), Cyclin F functions as a substrate receptor protein within the Skp1-Cullin-F-box E3 ubiquitin ligase complex, enabling the ubiquitylation of target proteins. The structural features that distinguish Cyclin F as a ligase adaptor and the mechanisms underlying its selective substrate recruitment over Cyclin A, which functions in complex with CDK2 at a similar time in the cell cycle, remain largely unexplored. We utilized single-particle cryoelectron microscopy to elucidate the structure of a Cyclin F-Skp1 complex bound to an E2F1 peptide. The structure and biochemical analysis reveal important differences in the substrate-binding site of Cyclin F compared to Cyclin A. Our findings expand on the canonical cyclin-binding motif (Cy or RxL) and highlight the importance of electrostatics at the E2F1 binding interface, which varies between Cyclin F and Cyclin A. These results advance our understanding of E2F1 regulation and may inform strategies for selectively targeting Cyclin F in cancer or neurodegeneration.
Li-Fraumeni syndrome (LFS) is a cancer predisposition syndrome caused by pathogenic/likely pathogenic germline TP53 variants. Core cancers include sarcomas, brain tumors, adrenocortical carcinoma and breast cancer. Surveillance with whole-body MRI … Li-Fraumeni syndrome (LFS) is a cancer predisposition syndrome caused by pathogenic/likely pathogenic germline TP53 variants. Core cancers include sarcomas, brain tumors, adrenocortical carcinoma and breast cancer. Surveillance with whole-body MRI (WBMRI) and other modalities is used for early cancer detection, regardless of the individual's personal cancer history. With increasing use of diagnostic multigene panels in oncology, more diverse phenotypic presentations have emerged, and subsequent cascade testing identifies more asymptomatic cancer-free individuals - 'previvors.' This review analyzes aspects of early cancer detection screening programs in asymptomatic germline TP53 variant carriers including current guidelines, specific founder variant populations, health economics and emerging strategies including liquid biopsies and wearable devices. A literature search with PubMed included publications in English until April 2025. Current guidelines recommend WBMRI in all LFS individuals, regardless of their prior cancer history due to its demonstrated survival advantage. Guidelines for use of other modalities such as endoscopy, ultrasound or laboratory tests are less well-established. Therefore, longitudinal prospective studies including all these modalities and their cancer detection rates are needed. In the future, a one-size-fits-all approach toward surveillance will be replaced by more precise patient-centered tailor-made screening programs incorporating noninvasive methods.
Abstract Background Li–Fraumeni syndrome (LFS) is a rare autosomal‐dominant cancer‐predisposition syndrome caused by germline pathogenic or likely pathogenic variants (P/LPVs) in the TP53 gene. Classical autosomal‐dominant inheritance predicts a 50% … Abstract Background Li–Fraumeni syndrome (LFS) is a rare autosomal‐dominant cancer‐predisposition syndrome caused by germline pathogenic or likely pathogenic variants (P/LPVs) in the TP53 gene. Classical autosomal‐dominant inheritance predicts a 50% transmission rate of TP53 P/LPV from each carrier parent to their offspring. However, clinical observations suggest higher‐than‐expected carrier proportions, indicating a potential transmission ratio distortion (TRD). The objective of this study was to investigate TRD in Israeli LFS families. Methods Data from families with an LFS diagnosis who were followed at Sheba Medical Center between 2015 and 2024 were reviewed. Families that had complete clinical data and offspring were included. Pedigree analyses classified carriers as confirmed, obligate, or probable. Observed carrier proportions were compared with the expected 50% rate using one‐sample t ‐tests. Results Among 171 individuals from 20 families, 100 (58.5%) were identified as TP53 P/LPV confirmed or obligatory carriers, significantly exceeding the expected 50% inheritance rate ( p = .027). A second analysis, which included 11 probable carriers, resulted in a carrier proportion of 64.9% ( p < .001). TRD was observed across all phenotypic groups. No significant differences in TRD were observed by sex or variant type. Conclusions This study revealed significant TRD in TP53 P/LPV inheritance among Israeli LFS families. A potential mechanism involves the role of TP53 in the cell cycle, in which reduced TP53 function may enhance embryonic cell proliferation, offering a survival or implantation advantage. TRD in LFS has implications for genetic counseling, reproductive decision making, and clinical management. These findings underscore the need for further research to validate TRD across diverse populations and elucidate the underlying mechanisms.
Chemotherapy for triple-negative breast cancer (TNBC) is often limited in efficacy due to drug resistance. The NOTCH1 pathway significantly contributes to the advancement of tumors, but its mechanism of action … Chemotherapy for triple-negative breast cancer (TNBC) is often limited in efficacy due to drug resistance. The NOTCH1 pathway significantly contributes to the advancement of tumors, but its mechanism of action in sensitizing TNBC to chemotherapy and its association with the downstream molecule, NT5E, is unclear. To explore the molecular mechanisms by which NOTCH1 regulates cisplatin sensitivity in TNBC cells, and to validate its synergistic effect with NT5E. Expression of NOTCH1 in MDA-MB-231 cells was silenced using RNA interference, and the changes in cell proliferation, migration and cisplatin sensitivity were measured in combination with cell function experiments. The regulatory relationship between NOTCH1 and NT5E was analyzed using qPCR and Western blotting, and the silencing effect of NOTCH1 was verified using NT5E overexpression experiments. Knockdown of NOTCH1 hindered the growth and motility of TNBC cells and lowered cisplatin's half-maximal inhibitory concentration. Expression of NOTCH1 and NT5E was positively correlated, and NOTCH1 silencing led to a decrease in the expression of NT5E. Elevated NT5E expression attenuated the suppressive effects of NOTCH1 knockdown on both cell proliferation and cisplatin response. NOTCH1 enhances TNBC cisplatin chemosensitivity by regulating NT5E expression. This study provides a new target and experimental basis for the development of combination therapy strategies for TNBC.
Abstract Li-Fraumeni syndrome (LFS) is a rare genetic disorder characterized by a predisposition to a variety of cancers due to the mutation in the protein TP53. The association between LFS … Abstract Li-Fraumeni syndrome (LFS) is a rare genetic disorder characterized by a predisposition to a variety of cancers due to the mutation in the protein TP53. The association between LFS and increased susceptibility to infections remains a currently underexplored area in the literature. In this report, we present 2 cases of patients with LFS who experienced both infectious and oncological complications due to their rare disorder. This includes a patient with persistent Enterococcus bacteremia, Aspergillus pneumonia, and refractory acute myeloid leukemia, and another patient with cellulitis unresponsive to antibiotics and stage IV adenocarcinoma. The main objective of these cases is to shed light on the variety of different cancers and infections associated with LFS and to highlight the importance of early recognition and aggressive treatment of infectious complications in LFS patients. Raising awareness of these topics is crucial to prevent the growth and spread of infections as a byproduct of oncologic treatments.
Abstract Butyrate-mediated inhibition of cell proliferation is part of the preventive role of dietary fiber against colorectal cancer (CRC). This effect notably involves the cyclin-dependent kinase inhibitor CDKN1A (p21 Cip/Waf1 … Abstract Butyrate-mediated inhibition of cell proliferation is part of the preventive role of dietary fiber against colorectal cancer (CRC). This effect notably involves the cyclin-dependent kinase inhibitor CDKN1A (p21 Cip/Waf1 ) in human intestinal cells, yet the underlying molecular mechanisms remain incompletely understood. Previously, we observed a paradoxical increase in cyclin D3 (CCND3)—but not cyclin D1—levels upon butyrate exposure. Here, we demonstrate that the butyrate-induced accumulation of CCND3 protein results both from mRNA increase and a CDKN1A-dependent protein stabilization, specifically extending its nuclear half-life. Proteomic analyses of CCND3 co-immunoprecipitates identified complexes involving CDKN1A, CDK4, CDK6, and the CRC-associated kinase CDK5, particularly enriched in butyrate-treated cells. Phosphorylation at a conserved Thr residue, crucial for CCND nuclear export and subsequent proteasomal degradation, was notably reduced following butyrate treatment and inversely correlated with CDKN1A expression levels. Structural modeling based on AlphaFold2, complemented by molecular dynamics simulations, revealed possible differential interactions between CDKN1A and cyclins D1 and D3, predicting that CCND3-Thr283 becomes structurally buried upon CDKN1A binding, limiting its phosphorylation. Our findings provide novel mechanistic insights into how CDKN1A might regulate CCND3 stability, highlighting previously unexplored roles of cyclin D3-containing complexes in cell cycle arrest induced by butyrate.
Protein phosphatase 1 regulatory subunit 12B (PPP1R12B) is a regulatory subunit of protein phosphatase 1. While our previous study identified the inhibitory role of PPP1R12B in hepatocellular carcinoma (HCC), the … Protein phosphatase 1 regulatory subunit 12B (PPP1R12B) is a regulatory subunit of protein phosphatase 1. While our previous study identified the inhibitory role of PPP1R12B in hepatocellular carcinoma (HCC), the precise molecular mechanisms underlying its anti-proliferative effects remain unclear. Herein, we demonstrated that PPP1R12B expression is significantly downregulated in HCC tissues and serves as an independent prognostic marker for favorable patient outcomes. Additionally, overexpression and silence of PPP1R12B experiments showed that PPP1R12B overexpression restricted cell proliferation and colony formation in vitro , and inhibited xenografted tumor growth in vivo , while its knockdown had opposite effects. Mechanistically, PPP1R12B could interact with p21-activated kinase 2 (PAK2) to suppress β-catenin expression and phosphorylation at Ser675, thereby impeding its nuclear translocation and subsequent transcriptional activation of Cyclin D1. This cascade culminated in G0/G1 phase cell cycle arrest. Furthermore, analysis of TCGA-HCC datasets confirmed inverse correlations between PPP1R12B and PAK2 or CTNNB1 (β-catenin) expression. Collectively, our findings elucidated a novel tumor-suppressive role of PPP1R12B in HCC through modulation of the PAK2/β-catenin/Cyclin D1 axis.
Background/Objectives: Oral squamous cell carcinoma (OSCC) is the most prevalent malignancy of the oral cavity and is frequently diagnosed at an advanced stage, resulting in poor prognosis and limited treatment … Background/Objectives: Oral squamous cell carcinoma (OSCC) is the most prevalent malignancy of the oral cavity and is frequently diagnosed at an advanced stage, resulting in poor prognosis and limited treatment options. Identifying reliable biomarkers that can predict tumor progression and serve as therapeutic targets remains an urgent clinical need. Methods: To identify key molecular drivers in OSCC, we performed an integrative bioinformatics analysis of five OSCC-related microarray datasets from the Gene Expression Omnibus (GEO). Differentially expressed genes (DEGs) were identified and subjected to functional enrichment, protein-protein interaction (PPI) network construction, and hub gene ranking using Cytoscape. Candidate genes were further validated using TCGA, UALCAN, and the Human Protein Atlas. In vitro functional assays were performed to evaluate the effect of TK1 knockdown on cell migration. Results: A total of 138 common DEGs were identified across datasets. GO enrichment revealed that these genes were associated with cell proliferation, extracellular matrix organization, and metastasis-related processes. Thymidine kinase 1 (TK1) was identified as a key hub gene and found to be consistently overexpressed in OSCC tissues. Kaplan-Meier analysis showed that high TK1 expression correlated with poor overall survival in head and neck cancer. TK1 knockdown in OSCC cell lines significantly impaired cell migration and wound-healing ability. Conclusions: Our findings suggest that TK1 plays an active role in promoting OSCC progression and may serve as a prognostic biomarker and potential therapeutic target for metastatic OSCC.
Abstract Gastrointestinal tract cancer is among the most common types of cancer and includes colorectal cancer (CRC) and gastric cancer (GC). Early tumor detection has been shown to reduce cancer‐associated … Abstract Gastrointestinal tract cancer is among the most common types of cancer and includes colorectal cancer (CRC) and gastric cancer (GC). Early tumor detection has been shown to reduce cancer‐associated mortality, emphasizing the significance of regular screening. In this regard, blood‐based tumor markers have gained popularity in cancer management. The purpose of this research was to examine the potential association between serum p53 antibodies and malignant tumors. A case–control study was conducted, including 37 GC samples, 53 CRC samples, and 62 healthy control samples. Serum levels of p53 antibodies were assessed using an enzyme‐linked immunosorbent assay. The results showed that the concentration of anti‐p53 antibodies in GC samples was 2.35 ng/mL, significantly ( p‐ value <.001) lower than the 9.24 ng/mL observed in the healthy control group. Similarly, the concentration of anti‐p53 antibodies in CRC samples was 4.14 ng/mL, which was significantly lower than the 9.31 ng/mL found in the healthy control group ( p‐ value <.001). These findings strongly suggest an association between the level of serum p53 antibodies and an elevated risk of cancer, implying their potential role as an early serological marker for the diagnosis of malignant tumors. However, it is important to note that measuring anti‐p53 antibodies alone may not be clinically effective in distinguishing CRC and/or GC from healthy controls. Further research and a comprehensive approach would be necessary for more accurate diagnostic outcomes.
Abstract Tissue homeostasis requires a precise balance between cellular self-renewal and differentiation. While fate decisions are known to be closely linked to cell cycle progression, the functional significance of this … Abstract Tissue homeostasis requires a precise balance between cellular self-renewal and differentiation. While fate decisions are known to be closely linked to cell cycle progression, the functional significance of this relationship is unclear. Here, we develop a mechanistic framework to analyse cellular dynamics when cell fate is coupled to cell cycle length. We focus on a distinct feature of cell cycle regulation where mitogens act as control parameters for a bifurcation governing the G1-S transition. Under competitive feedback from cell-cell interactions, the cell cycle regulatory network fine-tunes to the critical point of this bifurcation, becoming highly sensitive to mitogenic signalling. This critical positioning lengthens G1 while amplifying cell-to-cell variability in signalling and biochemical states. Such regulation confers significant advantages for controlling cell population dynamics, including maintaining a robust population set-point and rejecting mis-sensing mutants. The mutant rejection capability trades off against tissue growth and repair. Counter-intuitively, we propose that adult stem cells couple prolonged G1 with increased self-renewal propensity to efficiently eliminate mis-sensing mutants. Our theory explains and predicts regulatory patterns across development, homeostasis, and ageing.
ABSTRACT AXIN1 is a central regulatory hub of many oncogenic pathways in colorectal cancer (CRC). As the main scaffold protein and least abundant component of the beta-catenin destruction complex, changes … ABSTRACT AXIN1 is a central regulatory hub of many oncogenic pathways in colorectal cancer (CRC). As the main scaffold protein and least abundant component of the beta-catenin destruction complex, changes in AXIN1 levels affect Wnt signaling output. We show that targeting the Ras-MAPK pathway by MEK1/2 inhibitors induces AXIN1 loss across a panel of CRC cell lines and patient-derived organoids. GSK3B inhibition similarly reduced AXIN1 levels, yet by distinct mechanisms. MEK1/2 causes a reduction of AXIN1 transcript levels, but neither affects protein stability nor post-translational modifications of AXIN1. In contrast, GSK3B inhibition induces rapid AXIN1 degradation. Prevention of AXIN1 loss by co-treatment with tankyrase inhibitors was much stronger for GSK3B than for MEK1/2 inhibition. Using isogenic CRC cell lines and murine intestinal organoids, we show that APC truncations strongly reduce basal AXIN1 levels, but do not alter dynamics of AXIN1 loss upon MEK1/2 inhibition. Polysome profiling and Ribo-Seq revealed that MEK1/2 inhibition reduces global protein synthesis via an mTOR dependent pathway. This translational repression is sufficient to cause significant AXIN1 loss, as treatment with mTOR inhibitors phenocopies the effect of MEK1/2 inhibitors. Our study demonstrates that AXIN1 protein homeostasis is critically controlled by Ras-MAPK signaling at the level of protein synthesis, and that MEK1/2 inhibitors cause AXIN1 loss by translational repression.
Abstract Extensively studied over the past four decades, the TP53 gene has emerged as a pivotal watchman in cellular defense and a key factor in cancer biology. TP53 is the … Abstract Extensively studied over the past four decades, the TP53 gene has emerged as a pivotal watchman in cellular defense and a key factor in cancer biology. TP53 is the most frequently mutated gene in human malignancies, 50% of which carry alterations to it. Initially, the functions of p53 were thought to be restricted to cell‐cycle arrest and apoptosis. With time, however, a growing number of functions have been discovered, illustrating p53's role as a master switch between any cellular stress and cellular or multicellular responses that contribute to its anti‐tumor activity. Indeed, the peculiar landscape of TP53 mutations and its high heterogeneity are linked both to the structure of the protein and its ubiquitous function in regulating cellular homeostasis. Mutations in p53 are associated with poor response to therapy and shorter survival in most cancer types, and the diagnosis of p53 mutations is currently used to improve case management in some types of leukemia and lymphoma. Although TP53 has been defined as a tumor suppressor gene, overexpressed mutated p53 variants found in human tumors are defined as dominant oncogenes with a potential gain of function, which makes the gene a very attractive target for developing new cancer treatments. Beyond its role in cancer, our review also highlights TP53's significance in non‐neoplastic conditions, such as bone marrow failure syndromes and certain developmental disorders, where chronic p53 activation plays a crucial role in cellular stress responses, demonstrating its broader biological importance.
We have previously described a central role for CDK1 at the nexus of adhesion signalling and cell cycle progression, demonstrating that CDK1 has a non-canonical role in regulating integrin adhesion … We have previously described a central role for CDK1 at the nexus of adhesion signalling and cell cycle progression, demonstrating that CDK1 has a non-canonical role in regulating integrin adhesion complexes and in the migration of cancer cells in 3D interstitial matrix. Here we show that the CDK1 binding partners cyclinB1 and cyclinA2 also have roles in cell migration and invasion in both cancer and non-transformed cells. CyclinB1 plays a key role in RhoA activation to promote rear retraction in a membrane tension dependent manner, while cyclinA2 has a general role in promoting motility. Knockdown of either cyclin significantly perturbs migration with contrasting phenotypes, while knockdown of both together has an additive effect which arrests both migration and division. Our findings therefore describe how cyclin-CDK1 complexes orchestrate migration as well as division of cells and that cyclinA2-CDK1 and cyclinB1-CDK1 complexes play distinct roles in motility.
Abstract Approximately half of all cancers bear mutations in the tumor suppressor p53. Despite decades of research studying p53 function, treatment of p53-mutant cancers remains challenging owing to the effects … Abstract Approximately half of all cancers bear mutations in the tumor suppressor p53. Despite decades of research studying p53 function, treatment of p53-mutant cancers remains challenging owing to the effects of p53 mutations on many complex and interrelated signaling networks that promote tumor metastasis and chemoresistance. Mutations in p53 promote tumor survival by dysregulating cellular homeostasis and preventing activation of regulated cell death (RCD) pathways, which normally promote organismal health by eliminating dysregulated cells. Activation of RCD is a hallmark of effective cancer therapies, and p53-mutant cancers may be particularly susceptible to activation of certain RCD pathways. In this review, we discuss four RCD pathways that are the targets of emerging cancer therapeutics to treat p53-mutant cancers. These RCD pathways include E2F1-dependent apoptosis, necroptosis, mitochondrial permeability transition-driven necrosis, and ferroptosis. We discuss mechanisms of RCD activation, effects of p53 mutation on RCD activation, and current pharmaceutical strategies for RCD activation in p53-mutant cancers. Graphical Abstract
Abstract Background: Patients exhibiting exceptional response to targeted treatments represent a unique opportunity towards identifying predictive biomarkers of response. The introduction of edits into the genome using Clustered Regularly Interspaced … Abstract Background: Patients exhibiting exceptional response to targeted treatments represent a unique opportunity towards identifying predictive biomarkers of response. The introduction of edits into the genome using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) creates cancer models that can unravel the mechanisms driving tumorigenesis and response to drug agents. The study aimed to identify candidate molecular alterations associated with exceptional response to CDK inhibitors (CDKi) and validate their effects using genome editing. Methods: Patients with ER-positive, HER2-negative advanced breast cancer treated with CDKi and endocrine combination therapy at Departments of Oncology-affiliated with the Hellenic Cooperative Oncology Group (HeCOG), were included. Exceptional response was defined as either complete response (CR) to the treatment regimen or partial response (PR) for 3 years, per physician’s assessment. Formalin-fixed paraffin-embedded (FFPE) tumor tissue samples, obtained prior to CDKi administration from these patients were retrospectively collected. Whole-exome sequencing (WES) was performed to analyze tumor mutational landscape of exceptional responders to CDKi. Using CRISPR-mediated genome editing, a breast cancer cell model harboring a patient-specific SNP and the negative control were prepared, and subsequently evaluated for their responsiveness to palbociclib. Transcriptomic and proteomic analyses were conducted to uncover alterations in network interactions, between the two cell lines, elucidating the exceptional response to CDKi. Results: Overall, 12 women identified as exceptional responders, initiated CDKi treatment from March 2017 to February 2020; median age at diagnosis was 46 (range, 37 to 78). All patients had received the CDKi as first-line treatment; most commonly palbociclib (10 patients, 83.3%). Based on physician assessment, 50% of patients achieved CR with the combination therapy. After a median follow up of 65 months, 10 patients are still alive, and 7 are still receiving treatment with CDKi in combination with endocrine treatment. WES results with adequate mapped reads and acceptable technical characteristics in both tumor and control blood samples were obtained from 6 of 12 patients. A set of germline and somatic single nucleotide polymorphisms (SNPs) commonly shared in this cohort was identified. The focus was on MINDY1, a member of the MINDY family of deubiquitinases (DUBs), known as a deubiquitylase of ERα in breast cancer, the common SNP among all 6 patients. Utilizing CRISPR-based genome editing, a breast cancer cell model harboring a specific MINDY1 SNP was engineered, and their response to palbociclib was subsequently evaluated. Depletion of MINDY1 transcripts through RNA interference with short-hairpin RNA, increased sensitivity to palbociclib in both cell lines, confirming that MINDY1 protein is involved in molecular pathways related to palbociclib response. The functional role of MINDY1 rs771205 was also investigated by combining transcriptomic and proteomic data. Our data demonstrated that the MINDY1 rs771205 SNP, which causes an amino acid substitution, while it does not impact the respective transcript and protein levels, alters the MINDY1 protein’s interaction network. Conclusions: The Rs771205 single mutation alters the MINDY1 protein’s interaction network, sensitizing cells to palbociclib. Our study sheds new light on the underlying mechanisms driving exceptional response in patients with ER-positive, HER2-negative advanced breast cancer treated with CDKi and endocrine combination therapy , demonstrating that it is an opportune moment to invest attention and resources towards probing the underlying mechanisms of exceptional response in cancer patients. Citation Format: Eleni Balla, Yi Li, Kyriaki Papadopoulou, John T. Nguyen, Zikun Zhou, Anna Koumarianou, Eleni Galani, Rania Romanidou, George Fountzilas, Evangelia Razis, Angelos Koutras, Leonidas Bleris, Elena Fountzilas. MINDY1 role and exceptional response to CDK inhibitors in metastatic breast cancer [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P4-03-26.
MDM2 is a key negative regulator of the tumor suppressor p53 and an attractive target for cancer therapy. We report the discovery of MD-4251, the first orally efficacious MDM2 degrader … MDM2 is a key negative regulator of the tumor suppressor p53 and an attractive target for cancer therapy. We report the discovery of MD-4251, the first orally efficacious MDM2 degrader developed using PROTAC technology. MD-4251 induces potent and rapid MDM2 degradation in RS4;11 cells (DC50 = 0.2 nM; Dmax = 96% at 2 h), leading to robust p53 activation. It selectively inhibits the growth of acute leukemia cell lines with wild-type p53, with minimal activity in p53 mutant lines. MD-4251 shows excellent oral bioavailability in mice, favorable metabolic stability, and no CYP or hERG liabilities. A single oral dose induces sustained MDM2 depletion and attains complete tumor regression in vivo. These results support MD-4251 as a promising therapeutic candidate for cancers through depletion of MDM2.
Abstract The p53 mutation is the most common mutation in breast cancer, characterized by rapid proliferation, easy metastasis and poor prognosis. MAPK activated protein kinase 5, also named as PRAK, … Abstract The p53 mutation is the most common mutation in breast cancer, characterized by rapid proliferation, easy metastasis and poor prognosis. MAPK activated protein kinase 5, also named as PRAK, has been reported to function as either an oncogen or a tumor suppressor in different types of human cancers. The role of PRAK in breast cancer carcinogenesis and progression is still unknown. Our previous research found that PRAK function as a tumor suppressor in p53 wild breast cancer and function as an oncogene in p53 mutated breast cancer. In addition, PRAK influenced extracellular matrix (ECM) degradation and metastasis in breast by regulation of MMP9 expression. However, the molecular mechanism is still unknown. ChIP-seq combined with RNA-seq and protein profiling revealed that PRAK could bind to transcriptional factor CREB1 and wild p53. Moreover, we also observed the binding of PRAK to the MMP9 enhancer region. GST-pull down and IP were verified to bind from in vitro and in vivo; Transwell, scratch test and collagen contraction assay were used to evaluate the malignant phenotype of cells in vitro; inoculated SCID mice were observed to grow and metastasize in vivo by in vivo imaging; IHC was used to detect the protein expression of breast cancer tissues with different p53 statuses; and the correlation of its expression and its relationship to clinicopathological factors and prognosis were analyzed in the light of clinical data. In this study, we has found that in p53 wild-type breast cancer, PRAK binds to p53, and the activated p53 binds to the MMP9 promoter, which transcriptionally inhibits the expression of MMP9, prevents the degradation of extracellular matrix (ECM), and inhibits the metastasis of breast cancer; in p53 mutant breast cancer, PRAK binds to the MMP9 enhancer, CREB1 binds to the MMP9 promoter, and the combination of PRAK and CREB1 alters the conformation of the MMP9 genome, transcriptionally activates the expression of MMP9, induces the degradation of ECM, and promotes breast cancer metastasis. Ultimately, we elucidated the mechanism of PRAK in breast cancer with different p53 status and provide a theoretical basis for the development of therapeutic drugs in p53 mutated breast cancer. Citation Format: Jingjing Liu, Xu Liu, Shaorong Zhao, Jin Zhang. Dual function of PRAK in TP53-associated breast cancer metastasis [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P2-04-07.
The tumor suppressor p53 plays a crucial role in maintaining genome integrity in response to exogenous or endogenous stresses. The dynamics of p53 activation are stimulus- and cell type-dependent and … The tumor suppressor p53 plays a crucial role in maintaining genome integrity in response to exogenous or endogenous stresses. The dynamics of p53 activation are stimulus- and cell type-dependent and regulate cell fate. Acting as a transcription factor, p53 induces the expression of target genes involved in apoptosis, cell cycle arrest and DNA repair. However, transcription is not a deterministic process, but rather occurs in bursts of activity and promoters switch stochastically between ON and OFF states, resulting in substantial cell-to-cell variability. Here, we characterized how stimulus-dependent p53 dynamics are converted into specific gene regulation patterns by inducing diverse forms of DNA damage ranging from ionizing and UV radiation to clinically relevant chemotherapeutics. We employed single molecule fluorescence in-situ hybridization (smFISH) to quantify the activity of target gene promoters at the single-cell and single-molecule level. To analyse this comprehensive data set, we developed a new framework for determining parameters of stochastic gene expression by Bayesian inference. Using this combined theoretical and experimental approach, we revealed that features of promoter activity are differentially regulated depending on the target gene and the nature and extent of the DNA damage induced. Indeed, stimulus-specific stochastic gene expression is predominantly regulated by promoter activation and deactivation rates. Interestingly, we found that in many situations, transcriptional activity was uncoupled from the total amount of p53 and the fraction bound to DNA, highlighting that transcriptional regulation by p53 is a multi-dimensional process. Taken together, our study provides insights into p53-mediated transcriptional regulation as an example of a dynamic transcription factor that shapes the cellular response to DNA damage.
Abstract Background: Mutations in the TP53 gene occur in ∼51% of breast cancers and result in loss of p53 tumor suppressor function and tumor progression. Reactivation of wild-type p53 is … Abstract Background: Mutations in the TP53 gene occur in ∼51% of breast cancers and result in loss of p53 tumor suppressor function and tumor progression. Reactivation of wild-type p53 is an attractive therapeutic approach for breast cancers with a TP53 mutation including the more aggressive triple-negative breast cancer (TNBC), where treatment options are limited. Rezatapopt (also known as PC14586) is an investigational first-in-class, p53 reactivator that selectively binds to the mutated p53 Y220C protein and stabilizes the structure in wild-type conformation, thereby restoring p53 wild-type activity. PYNNACLE (NCT04585750) is a Phase 1/2 clinical trial of rezatapopt in patients with locally advanced or metastatic solid tumors harboring the TP53 Y220C mutation. In Phase 1, rezatapopt demonstrated favorable safety and anti-tumor activity in heavily pre-treated patients (n=67 treated at the efficacious dose range of 1150 mg once daily [QD] to 1500 mg twice daily [BID]). This subgroup analysis assessed rezatapopt in patients with advanced breast cancer treated across the efficacious dose range. Methods: Patients with locally advanced or metastatic breast cancer with a TP53 Y220C mutation were eligible to receive rezatapopt orally on a continuous schedule across the efficacious dose range (1150 mg QD to 1500 mg BID). Safety and preliminary efficacy, as evaluated by the investigator using Response Evaluation Criteria in Solid Tumors (RECIST) v1.1, were assessed. Tumor next-generation sequencing was used to determine TP53 Y220C, BRCA, PIK3CA, and KRAS tumor mutation status. Results: As of September 5, 2023, nine patients with breast cancer (HR+/HER2- n=3; HR+/HER2+ n=1; HR-/HER2+ n=1; TNBC n=4) received rezatapopt in the PYNNACLE Phase 1 trial. The median age was 53 years (range 32–65 years) and Eastern Cooperative Oncology Group (ECOG) performance score was 0 (n=3) or 1 (n=6). One patient had a somatic BRCA2 mutation, no patient had a BRCA1 mutation, two patients had a PIK3CA mutation, and all patients were KRAS wild-type. The median number of prior lines of systemic therapy was 4 (range 2–9). Of the eight patients who had measurable disease at baseline, three (37.5%) achieved a confirmed partial response (PR), four had stable disease (SD), and one had progressive disease (PD) as the best objective response. Reductions in target lesions were reported in all patients, with a maximum reduction in tumor volume from baseline ranging from -2.4% to -55.2%. The most frequent treatment-related adverse events in patients with breast cancer were consistent with the overall Phase 1 PYNNACLE population with solid tumors treated at the efficacious dose range (n=67), and included nausea (51%), vomiting (43%), and increased blood creatinine (27%), which were mostly grade 1/2. Administration of rezatapopt with food led to an improvement in gastrointestinal adverse events including nausea and vomiting. Conclusions: In this Phase 1 PYNNACLE trial, rezatapopt demonstrated promising preliminary single-agent efficacy in heavily pre-treated patients with advanced breast cancer, including TNBC, harboring a TP53 Y220C mutation. Rezatapopt had a favorable safety profile in the efficacious dose range with improvements in gastrointestinal adverse events observed when administered with food. The PYNNACLE tumor-agnostic registrational Phase 2 trial, which includes a breast cancer cohort, will assess rezatapopt as monotherapy at the recommended Phase 2 dose of 2000 mg QD with food in patients with TP53 Y220C-mutated and KRAS wild-type advanced solid tumors. Citation Format: Ecaterina Dumbrava, Shivaani Kummar, Melissa Johnson, Kim Le Duke, Yajuan G Qin, Marc Fellous, Alison M Schram. Phase 1 Analysis from the PYNNACLE Phase 1/2 Study of Rezatapopt in the Subgroup of Patients with Advanced Breast Cancer Harboring a TP53 Y220C Mutation [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P3-12-27.
Abstract Background: Multiple studies has shown that HOXB13 gene expression is associated with poor prognosis for hormone receptor positive (HR+) post-menopausal breast cancer (BC) patients. However, the biological contribution of … Abstract Background: Multiple studies has shown that HOXB13 gene expression is associated with poor prognosis for hormone receptor positive (HR+) post-menopausal breast cancer (BC) patients. However, the biological contribution of HOXB13 in such patients is unclear. Previous gene expression- and proteomic-based analyses of HOXB13-high and -low human breast cancers and breast cancer cell lines indicate that HOXB13 expression is associated with interferon gene signatures, suggesting that HOXB13 may modulate the tumor immune microenvironment (TIME). Herein, we investigate the role of HOXB13 expression in breast cancer development in two independent mouse mammary tumor transplant models. Methods: Murine HOXB13 was ectopically expressed the ER+ SSM3 and the ER- PyMT mouse mammary tumor cell lines, and these cells along with non-HOXB13 expressing control cell lines were transplanted into the inguinal mammary fat pad of syngeneic wild-type (WT) mice on the 129S6/SvEv and C57BL/6 backgrounds, respectively. Tumor volumes and weight were monitored every other day and mice sacrificed at day 40. Tumor single-cell suspensions were prepared, and tumor infiltrating lymphocytes (TILs) were isolated using CD45+ Microbeads on magnetic columns. Cells were stained with the following surface antibodies: CD3, CD4, CD8α. After fixation and permeabilization, intracellular stains were performed using antibodies to TNF-a and TGF-b. Stained cells were assayed using a BD LSRFortessa flow cytometer, and data were analyzed using FlowJo software. To demonstrate that mature T and/or B cells lymphocytes play an important role in the modulation of tumor growth, PyMT-HOXB13 and -control cells were transplanted into Rag1 knockout (Rag1 KO) mice on the C57BL/6 background. Results: The SSM3-HOXB13 and PyMT-HOXB13 cells exhibited significantly increased tumor growth rate compared with their corresponding control cells after implantation into the inguinal mammary fat pad of WT 129S6/SvEv and C57BL/6 mice, respectively (Figure 2A). Analysis of tumor-infiltrating lymphocytes revealed a marked decrease in CD3+ T and CD8+ T cells in both the ER+ SSM3-HOXB13 and the ER- PyMT-HOXB13 tumors, along with a significantly reduced percentage of activated (TNF-α+ and IFN-γ+) CD8+ T cells in these tumors. Importantly, PyMT-HOXB13 and PyMT-Ctrl tumors grew at a similar rate in the inguinal mammary fat pad of Rag1 KO mice lacking T and B lymphocytes. Conclusions: HOXB13 expression in two independent mouse mammary tumor models confers tumoral growth advantage by impeding antitumor T cell immunity. Citation Format: Dennis Sgroi, Yun Xia, Marinko Sremac, Shawn Demehri. HOXB13 Expression Induces an Immunosuppressive Tumor Microenvironment in Breast Cancer [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P2-06-13.
Abstract Background: Triple-negative breast cancer (TNBC) has the poorest prognosis of the breast cancer subtypes, in part due to the lack of targeted therapy options. TNBCs lack the expression of … Abstract Background: Triple-negative breast cancer (TNBC) has the poorest prognosis of the breast cancer subtypes, in part due to the lack of targeted therapy options. TNBCs lack the expression of targetable cell surface or nuclear receptors such as estrogen receptor (ER) or HER2 present in other breast cancers. Standard of care for TNBC patients includes surgery, radiation, and non-specific chemotherapy, and very few women with TNBC meet criteria to receive immunotherapy or PARP inhibitors. Thus, new targeted therapies are urgently needed for these aggressive cancers. Nearly all TNBCs harbor TP53 mutations, so our laboratory has sought to identify molecular vulnerabilities of these TP53-mutant breast cancers. A drug screen identified that the Kif11 inhibitor SB-743921 preferentially killed TP53 mutant breast cancer cells (as compared to TP53-wild type cells). Kif11 is a motor protein critical for proper alignment of the mitotic spindle, and inhibition leads to mitotic defects and cell cycle arrest. Hypothesis: We hypothesized that Kif11 inhibition causes TP53-mutant TNBC cells to undergo mitotic catastrophe leading to cell death. Methods: Clinical Data: Data was accessed via cBioPortal or UCSD Xena Browser. Expression was compared between groups using Student’s t-tests. For Kaplan-Meier curves, Kif11 expression was dichotomized at the median and survival curves were compared using the Log-Rank test. Cell Proliferation and Cell Death Assays: Cells were treated with SB-743921 or vehicle, stained with Hoechst 33342 and DRAQ7(2D) or Annexin V-FITC and PI (Flow), and imaged on the ImageXpress PICO or analyzed via flow cytometry. Immunofluorescence: Cells were treated with SB-743921 or vehicle, stained with anti-alpha tubulin-AlexaFluor488 and DAPI and imaged on the Nikon Ti2. Mouse xenograft experiments: Cells were injected into the mammary fat pads of nude mice. When tumors reached 50-100 mm3, mice were treated with the Kif11 inhibitor SB-743921 10mg/kg or vehicle i.p. 3 times weekly. Experiments were conducted under IACUC approval. Results: The Kif11 mitotic kinesin is more highly expressed in TP53 mutant and TNBCs than in TP53 wild-type breast cancers, and high expression is associated with poorer survival. Kif11 inhibition leads to cell cycle arrest and growth suppression in both TP53-wild-type and TP53-mutant breast cancer cells. However, in TP53-mutant cells Kif11 inhibition causes mitotic dysfunction, formation of multinucleated giant cells, and caspase-dependent apoptotic cell death. Furthermore, knockout of TP53 in wild-type breast cancer cells sensitizes them to cell death following Kif11 inhibition. Kif11 inhibition also inhibits the growth of TP53-mutant TNBC xenografts in vivo. Conclusions: In cells with TP53-mutation or loss, Kif11 inhibition results in mitotic dysfunction and cell death due to mitotic catastrophe, and Kif11 inhibitors suppress in vivo TP53-mutant TNBC growth. These results support further investigation of Kif11 inhibitors as a potential therapeutic in TP53 mutant TNBC. Acknowledgments: This work was supported by the John Charles Cain Endowment, Pauline Altman Goldstein Discovery Research Award, and the NCATS of the NIH under Award Numbers TL1TR003169 and UL1TR003167. We also thank the MDACC NORTH Campus Flow Cytometry and Cellular Imaging Core Facility for their assistance. Citation Format: Amanda Lanier, William Tahaney, Cassandra Moyer, Jamal Hill, Darrian Coleman, Banu Arun, Abhijit Mazumdar, Powel H Brown. Inhibition of the Kif11 mitotic protein kills TP53-mutant triple-negative breast cancer cells [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P2-02-10.
Abstract The paradigm of modern cytotoxic chemotherapy is to impair DNA replication to prevent cell division. During this process, newly synthesized DNA is wrapped around histones produced de novo to … Abstract The paradigm of modern cytotoxic chemotherapy is to impair DNA replication to prevent cell division. During this process, newly synthesized DNA is wrapped around histones produced de novo to create the fundamental unit of chromatin: nucleosomes. Yet, despite the 50+ approved oncology drugs that target DNA synthesis, there are currently no therapeutic strategies that target nucleosome production. This gap represents a large and untapped opportunity to develop novel, safer and more efficacious anti-cancer therapies. We have focused on histone acetyltransferase 1 (HAT1), an enzyme that acetylates and stabilizes newly synthesized histones in the cytosol to support nucleosome production. Our previously published data demonstrated that HAT1 controls histone production by also binding histone H4 promoters (Gruber et al. Mol. Cell 2019). In the absence of HAT1, cells cannot effectively synthesize nucleosomes and tumor growth is impaired. To further develop rationale for targeting HAT1 for breast cancer we bred the HAT1+/-heterozygous mouse to the MMTV-PyMT model of triple-negative breast cancer. Despite normal development, animals deficient for a single HAT1 allele in the MMTV-PyMT background evinced delayed mammary tumor onset, decreased tumor size and increased survival compared to littermate controls (median survival 172 v. 196 days for WT v. het; log-rank p < 0.004). Histologic examination of tumors revealed a strong correlation between Ki-67 and HAT1 protein expression (R2 = 0.51, p < 0.01). HAT1 expression in heterozygous animals was primarily diminished in normal mammary glandular structures compared to WT controls, but HAT1 levels were high in tumors, which mirrored high expression of HAT1 and related co-factors in a panel of human triple-negative breast cancers. Since HAT1 primarily binds and regulates histone H4 promoters to coordinate nucleosome production further experiments were performed to test the requirement for histone H4 genes for mammary tumorigenesis. Biallelic CRISPR/Cas9 editing of either promoter or coding regions of three separate histone H4 genes in triple-negative breast cancer cell lines demonstrated that all edits diminished tumor growth potential in vivo. In addition, tumor growth could be rescued by re-expressing a single copy of histone H4 into cells bearing H4 gene edits, demonstrating the specificity of these edits. These results establish that the HAT1-histone H4 axis is a critical dependency for triple-negative breast cancer growth. Our lab has recently published the first small molecule HAT1 enzymatic inhibitor, which demonstrated anti-tumor activity and animal safety in vivo (Gaddameedi et al. J. Med. Chem. 2023). Thus, small molecule strategies to target the HAT1-H4 axis could have therapeutic relevance for triple-negative breast cancer with a goal of improving patient outcomes. Citation Format: Joshua Gruber, Harsh Goar, Shreenidhi Rajkumar, Danielle Dixon. Targeting nucleosome synthesis as a therapeutic strategy for triple-negative breast cancer [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P3-04-24.
Abstract In a normal cell cycle, there is redundancy in the role of the cell-cycle dependent kinases (CDKs) in regulating G1/S phase transition. In cancer cells, the regulation of G1/S … Abstract In a normal cell cycle, there is redundancy in the role of the cell-cycle dependent kinases (CDKs) in regulating G1/S phase transition. In cancer cells, the regulation of G1/S transition can be subverted by (a) amplification and elevated expression of Cyclin E (CCNE), or (b) mutation/loss of the Retinoblastoma 1 (RB1) gene. Cancer cells with these genomic alterations have been shown to exhibit profound sensitivity to CDK2 depletion, validating CDK2 as a potential therapeutic target. Here, we report the discovery and preclinical characterization of ETX-197, a highly potent and selective small molecule inhibitor of CDK2 enzymatic activity. Based on known ligand-CDK2 structures, ETX-197 is designed to induce previously unexplored interactions within the CDK2 ATP binding pocket leading to improved potency and selectivity compared to other known CDK2 inhibitors. ETX-197 is >100-fold selective against other kinases in the CDK family and the selectivity extends more broadly against 385 other kinases. The affinity of ETX-197 for CDK2 results in tight binding (slow off-rate) and high potency in pharmacodynamic modulation and anti-proliferative activity in vitro and in vivo. Treatment of CCNE-amplified cancer cells with ETX-197 results in concentration-dependent inhibition of pRB phosphorylation, G1/S phase cell-cycle arrest and cell proliferation. In addition, ETX-197 treatment phenocopies CDK2 genetic knock-down in cells, as revealed by bulk-RNA Seq analysis of CCNE-amplified or wild-type cells, confirming that the cellular activity of ETX-197 is on-target and highly selective. In mouse xenograft studies using CCNE-amplified ovarian cancer cell line (OVCAR-3) or patient-derived tumors, ETX-197 treatment causes dose-dependent tumor growth inhibition with excellent tolerability. Interestingly, in RB1-deficient small cell lung cancer cell lines, ETX-197 treatment results in G2/M cell cycle arrest, accumulation of DNA damage, and apoptosis. Xenograft studies with small cell lung cancer cell lines and patient-derived tumor cells also show significant tumor growth inhibition with ETX-197. Additionally, ETX-197 has single-agent efficacy in a breast cancer xenograft model that had acquired resistance to a CDK4/6 inhibitor. These data suggest that ETX-197 has the potential to be a best-in class CDK2 inhibitor for the treatment of cancer with CCNE amplification or RB1 deficiency, including breast cancer that has progressed on treatment with a CDK4/6 inhibitor because of these genomic alterations. Currently, ETX-197 is being clinically developed by BeiGene in a first-in-human (FIH), Phase 1a/1b study to assess the safety, tolerability, pharmacokinetics (PK), pharmacodynamics, and preliminary antitumor activity in patients with advanced, nonresectable, or metastatic solid tumors (NCT06257264). Citation Format: Daliya Banerjee, Alexandra Weinheimer, Jingyan Gao, Fei Pang, Ying Lin, Raj Nagaraja, Yong Tang, Zipeng Fan, Zipeng Fan, Minghong Hao, Shengfang Jin, Tao Liu, Tai Wong. ETX-197/BG-68501, a potential best-in-class potent, selective, oral, small molecule CDK2 inhibitor, has anti-tumor activity in cancer models with Cyclin E amplification or deficiency in the Retinoblastoma 1 gene [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P4-12-29.
Abstract The single-stranded DNA cytosine deaminase enzyme, APOBEC3B (A3B), has been implicated as a mutational driver in multiple human cancers1,2. Breast cancer in particular shows high levels of A3B expression … Abstract The single-stranded DNA cytosine deaminase enzyme, APOBEC3B (A3B), has been implicated as a mutational driver in multiple human cancers1,2. Breast cancer in particular shows high levels of A3B expression and positive associations with poor clinical outcomes3. Recent studies have also demonstrated that human A3B expression in mice is carcinogenic with significantly shortened life expectancies4. The only genetic factor in breast cancer thus far associated with A3B expression and APOBEC signature single base substitutions is p53 mutation5. Therefore, here we test the hypothesis in vivo that human A3B expression will show pathlogical synergy in the form of accelerated mammary tumor penetrance upon combination with p53 haploinsufficiency due to a heterozygous deletion mutation spanning exons 2-10. Custom and commercially available animals were subjected to standard breeding practices to generate experimental (A3B, p53Delta exons 2-10/+) and control groups (p53Deltanexons 2-10/+). Surprisingly, in contrast to A3B-accelerated tumor development on a wildtype genetic background, A3B had no effect on the rates of tumor development in p53 heterozygous animals (median 13.5 months). The observed genetic epistasis was not due to a lack of A3B expression or activity, as evidenced by strong IHC positivity and a clear acculumlation of APOBEC signature single base substitution mutation in tumors. We conclude that A3B-accelerated tumor development requires full p53 function, and that a haploinsufficiency in p53 enables tumor cells (or their precursors) to better tolerate DNA damage lesions induced by A3B. Selected References: 1. Petljak, M. et al. Addressing the benefits of inhibiting APOBEC3-dependent mutagenesis in cancer. Nat Genet 54, 1599-1608 (2022). 2. Butler, K. & Banday, A. R. APOBEC3-mediated mutagenesis in cancer: causes, clinical significance and therapeutic potential. J Hematol Oncol 16, 31 (2023). 3. Roelofs, P. A. et al. Clinical implications of APOBEC3-mediated mutagenesis in breast cancer. Clin Cancer Res 29, 1658-1669 (2023). 4. Durfee, C. et al. Human APOBEC3B promotes tumor development in vivo including signature mutations and metastases. Cell Rep Med 4, 101211 (2023). 5. Burns, M. B. et al. APOBEC3B is an enzymatic source of mutation in breast cancer. Nature 494, 366–370 (2013).\ Citation Format: Joshua Proehl, Cameron Durfee, Yuan Zhao, Nuri Alpay Temiz, Reuben Harris. p53 Haploinsufficiency Epistatically Masks A3B Tumor Phenotype in vivo [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P3-06-26.
Abstract Introduction: TP53 is a multi-functional tumor suppressor gene, with several important roles in tumorigenesis. Females who harbor germline TP53 pathogenic/likely pathogenic variants (GPV) have a very high lifetime risk … Abstract Introduction: TP53 is a multi-functional tumor suppressor gene, with several important roles in tumorigenesis. Females who harbor germline TP53 pathogenic/likely pathogenic variants (GPV) have a very high lifetime risk of developing breast cancer (BC), especially hormone receptor positive and HER2 positive tumors. Specific TP53 variants have different functional consequences for molecular pathways and somatic aberrations in cancers, leading to the hypothesis that the type of TP53 GPV could modulate breast tumor phenotype. Objective: To examine differences in TP53 GPV and their predicted functional impact on age at first BC diagnosis and BC subtype (ER and HER2 status). MATERIAL AND METHODS: This multicenter international cohort was comprised of female TP53 GPV carriers diagnosed with any invasive BC (non-metastatic or de novo metastatic). We included TP53 variants classified as GPV in ClinVar, or classified as GPV by at least one major laboratory, or truncating variants. Carriers of a second GPV in other BC genes and carriers of TP53 GPV downgraded to a variant of uncertain significance by the TP53 ClinGen-VCEP were excluded. Clinical data were abstracted from medical records. TP53 GPV were classified according to mutation type: nonsense, frameshift, missense affecting the DNA binding domain (Missense_DBD) or the tetramerization domain (Missense_TD), variants affecting splicing, and copy number variations. For this report, functional classification was performed using Fortuno’s classification based on two germline TP53 databases from commercial labs. Functional classification categories included: missense variants with dominant negative effect (Missense_DNE) and without DNE (Missense_notDNE), truncating and hotspots variants. The bivariate association between tumor phenotype and each of the variables (mutation type and Fortuno’s functional classification) was assessed using two-sample Wilcoxon test for continuous and Fisher’s exact test for nominal categorical variables. Results: Among 301 females who met study criteria,mean age of BC diagnosis was 37.0 years (SD 10.46), 187 (62.1%) met Li-Fraumeni syndrome clinical criteria (revised Chompret or Classic criteria), and 41 (13.6%) had bilateral synchronous BC. The distribution of BC subtypes was: 108 (35.9%) ER+/HER2-, 79 (26.2%) ER+/HER2+, 62 (20.6%) ER-/HER2+, 20 (6.6%) ER-/HER2-, and 32 (10.6%) ER unknown and/or HER2 unknown. Most TP53 GPV were missense variants (n=217; 141 Missense_DBD, 76 Missense_TD). In comparison to Missense_TD, Missense_DBD carriers had a younger age at BC diagnosis (35.7 vs 42.3, p<0.01), lower rates of ER+ disease (63.1% vs 80.3%, p<0.01), and higher rates of HER2+ disease (48.9% vs 27.6%, p<0.01). When considering ER and HER2 subtypes, Missense_TD carriers had more ER+/HER2- BC (51.3% vs 34.8%, p=0.02) and Missense_DBD carriers had more ER-/HER2+ tumors (22.0% vs 5.3%, p=<0.01). According to Fortuno’s classification, 89 (29.6%) TP53 GPV were Missense_notDNE, 59 (19.6%) Missense_DNE, 80 (26.6%) truncating, 49 (16.3%) hotspots variants, and 24 (8.0%) variants unclassified by the method. Among all these categories, Missense_notDNE variants were associated with older ages of BC diagnosis (51.7% above 40 years; p<0.001), higher rates of ER+ disease (79.8%; p<0.001), lower rates of HER2+ tumors (29.2%; p<0.001), and lower rates of bilateral synchronous BC (p=0.003). CONCLUSIONS: These findings suggest that TP53 GPV functional status can influence age at breast cancer presentation and tumor ER/HER2 status which can potentially improve targeted treatment strategies and inform risk prediction and risk reduction strategies. Citation Format: Renata L. Sandoval, Michele Bottosso, Miki Horiguchi, Natalia Polidorio, Anh Le, Brittany L. Bychkovsky, Benjamin Verret, 
Alessandra Gennari, Sophie Cahill, Alison Schwartz-Levine, Olivier Caron, Marion ImbertBouteille,
Catherine Noguès, Pauline Rochefort, Kara N. Mawell, Maria Isabel Achatz, Fabrice Andre, Judy E. Garber. Differences in breast cancer phenotype by germline TP53 variant functional classification [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr PS16-04.
Backround Feline oral squamous cell carcinoma (FOSCC) is the most common malignant oral tumor in cats, characterized by invasive and aggressive behavior regardless of its location. Conventional treatments, including surgery, … Backround Feline oral squamous cell carcinoma (FOSCC) is the most common malignant oral tumor in cats, characterized by invasive and aggressive behavior regardless of its location. Conventional treatments, including surgery, radiation therapy, and chemotherapy, often yield unsatisfactory outcomes, with tumor progression and tissue destruction frequently leading to euthanasia. In anthroposophical medicine, extracts of Viscum album have been developed as complementary cancer treatments, with Iscador, the oldest and most widely used oncological drug, showing promising anticancer potential. This study investigated, for the first time, the cytotoxic and apoptotic effects of IscM and IscQu, two Viscum album extracts, on FOSCC cells. Methods Using primary cultures of three FOSCC cell lines, cell viability assays were performed to assess cytotoxicity, and the effects on apoptotic cell death, cell cycle arrest, and cellular and nuclear morphology were evaluated. Additionally, mRNA expression levels of Cyclin D, Cdk4, Bcl-2, Bax, and p53 were analyzed. Results The results revealed that both IscM and IscQu induced apoptotic cell death and promoted cell cycle arrest in all three FOSCC cell lines tested. IscQu exhibited relatively stronger pro-apoptotic effects compared to IscM, although no significant differences were observed among the cell lines. Conclusion These findings suggest that Viscum album extracts, particularly IscQu, may exert anti-tumor effects on feline oral squamous cell carcinoma cells in vitro .
Summary Colorectal carcinoma (CRC) is characterized by mutations in Wnt signaling pathway components and the p53 protein, and anti□PD-1/anti□PD-L1 immunotherapy has shown limited efficacy in microsatellite stable CRC patients. In … Summary Colorectal carcinoma (CRC) is characterized by mutations in Wnt signaling pathway components and the p53 protein, and anti□PD-1/anti□PD-L1 immunotherapy has shown limited efficacy in microsatellite stable CRC patients. In this work, CEABP1, a de novo-designed binding protein for the CRC marker carcinoembryonic antigen (CEA), and cell-penetrating peptide (CPP) were employed for targeted delivery in CRC cells. The consensus binding motifs for TCF/LEF and Max were concatenated, and Max DNA binding domain protein fused with CPP and CEABP1 was used to deliver this TCF/LEF transcription factor decoy (TFD) DNA specifically into CRC cells by recognizing CEA on the CRC cell surface to inhibit Wnt target gene transcription, leading to marked suppression of CRC cell proliferation and xenograft tumor growth. In addition, p28 was employed as a CPP to specifically deliver the p28-p53-CEABP1 protein into CRC cells, which significantly enhanced p53 inhibition of CRC cell proliferation and xenograft tumor growth. Furthermore, codelivery of the p14 ARF protein together with p53 increased its antitumor activity by prolonging its effective duration. These findings paved the way for the development of related biomacromolecular anticancer therapeutics.
Ovarian cancer is the sixth leading cause of cancer death among American women, with most fatalities attributable to tubo-ovarian high-grade serous carcinoma (HGSC). This malignancy usually develops resistance to conventional … Ovarian cancer is the sixth leading cause of cancer death among American women, with most fatalities attributable to tubo-ovarian high-grade serous carcinoma (HGSC). This malignancy usually develops resistance to conventional chemotherapy, underscoring the need for robust preclinical models to guide the development of novel therapies. Here, we introduce an HGSC mouse model generated via Ovgp1 -driven Cre recombinase effecting CRISPR/Cas9-mediated deletion of Trp53, Rb1 , and Nf1 tumor suppressors in mouse oviductal epithelium ( m-sgPRN model). Cyclin-dependent kinase 12 (CDK12) inactivation—frequently observed in human HGSC—is associated with poorer outcomes, DNA damage accumulation (including tandem duplications), and increased tumor immunogenicity. In our system, coablation of Cdk12 ( m-sgPRN;Cdk12KO ) recapitulated hallmark features of HGSC, while accelerating tumor progression and reducing survival. In a conventional (Cre-lox-mediated) Trp53/Nf1/Rb1 triple knockout model with concurrent Cdk12 ablation ( PRN ; Cdk12KO mice), we observed T cell–rich immune infiltrates mirroring those seen clinically. We established both models as subcutaneous or intraperitoneal syngeneic allografts of CDK12 -inactivated HGSC that exhibited sensitivity to immune checkpoint blockade. Furthermore, a CRISPR/Cas9 synthetic lethality screen in PRN;Cdk12KO -derived cell lines identified CDK13—an essential paralog of CDK12—as the most depleted candidate, confirming a previously reported synthetic lethal interaction. Pharmacologic CDK13/12 degradation (employing YJ1206) demonstrated enhanced efficacy in cell lines derived from both m-sgPRN;Cdk12KO and PRN ; Cdk12KO models. Our results define CDK12 as a key tumor suppressor in tubo-ovarian HGSC and highlight CDK13 targeting as a promising therapeutic approach in CDK12 -inactive disease. Additionally, we have established valuable in vivo resources to facilitate further investigation and drug development in this challenging malignancy.
Abstract Metastasis is a major cause of cancer deaths, but the underlying molecular mechanisms remain largely unknown. Esophageal squamous cell carcinoma (ESCC) is a highly aggressive cancer with poor survival, … Abstract Metastasis is a major cause of cancer deaths, but the underlying molecular mechanisms remain largely unknown. Esophageal squamous cell carcinoma (ESCC) is a highly aggressive cancer with poor survival, yet the key kinases driving ESCC metastasis and their biological function have not been fully discovered. Here, a kinase‐substrate map of metastatic ESCC is presented for the first time by conducting a phosphoproteomics analysis of 60 clinical specimens. By further consolidating data with CRISPR/Cas9 functional screening, LIM domain kinase 1 (LIMK1) is identified as a novel kinase of β‐catenin. The in vitro and in vivo experiments demonstrated that LIMK1 cooperates with Cyclin‐dependent kinase 5 (CDK5) to promote cancer metastasis in a phosphorylation‐dependent manner. Mechanistically, LIMK1 and CDK5 synergistically phosphorylate β‐catenin at S191, enhancing its phosphorylation and interaction with Nucleoporin 93, resulting in β‐catenin nuclear translocation and activation of key pathways in cancer metastasis. High expression of LIMK1 and CDK5 is associated with poor prognosis of ESCC patients, and the clinical and functional significance of LIMK1/CDK5‐Wnt/β‐catenin axis is also verified in esophageal adenocarcinoma, gastric cancer, and lung cancer. Furthermore, the combination of LIMK1 and CDK5 inhibitors significantly suppresses metastasis in multiple models. This work highlights LIMK1 as a novel regulatory and targetable kinase of β‐catenin, informing the treatment of advanced cancer.