High-grade serous ovarian carcinoma (HGSOC), which accounts for approximately 75% of ovarian cancer cases, is associated with poor clinical outcome. Although most patients initially achieve a complete response to conventional chemotherapy, HGSOC almost invariably develops chemoresistance. There is therefore an urgent need to identify predictive biomarkers of treatment response. Here, through integrative analyses of molecular and clinical data from HGSOC patient cohorts, we identify syntabulin (SYBU), a microtubule-associated protein originally described as a regulator of mitochondrial transport along neuronal microtubules, as a critical determinant of chemosensitivity in HGSOC. Low SYBU expression in tumors correlates with higher tumor grade and increased aggressiveness, yet paradoxically with enhanced sensitivity to chemotherapy. SYBU-deficient cancer cells display impaired oxidative phosphorylation and a metabolic shift toward glycolysis characteristic of the Warburg effect, together with mitotic defects such as chromosome lagging that promote aneuploidy. Mechanistically, syntabulin forms a complex with the mitochondrial outer membrane porin VDAC1 and the inner membrane protein MIC60, a major regulator of mitochondrial cristae organization. Functionally, the syntabulin-MIC60 axis controls cristae architecture and mitotic fidelity, thereby connecting mitochondrial metabolism to cell division. These findings highlight new therapeutic vulnerabilities to overcome chemoresistance in ovarian cancer. ### Competing Interest Statement The authors have declared no competing interest. Fondation ARC pour la Recherche sur le Cancer, https://ror.org/0489qz649, Post-doctoral fellowship for H.M, Subvention 2017-2019 for C.N, Passerelle Grant for E.P, MetaboPlast-ARCPJA2022060005283 for C.B Fondation de France, 00119147/WB-2021-35655 for H.M La Ligue Contre le Cancer, https://ror.org/00rkrv905, PhD Fellowship for M.M, Ligue Contre le Cancer 94/Val de Marne subvention for C.N Entreprises contre le Cancer Paris GEFLUC, metabolic analysis for G.G and F.M.-G, GEFLUC SYBU subvention 2024 for C.N, GEFLUC SYBU subvention 2025 for C.N, GEFLUC subvention 2023-2024 for C.B Institut National du Cancer (INCa), INCa 2017-1-PL BIO-08 and 2021 - 167/ INCA_16344 for C.B Société française de lutte contre les cancers et les leucémies de l’enfant et de l’adolescent (SFCE), ECS 20, PHC PESSOA, N°49163TL for C.B Eva pour la vie-Grandir Sans Cancer, EPLVGSD2025-Brenner for C.B Fondation Rothschild, Paris, France, Subvention Ovarian cancer chemoresistance 2023-2025 for C.N University Paris Saclay, Emergence LERMIT MitoMicro 2021 for C.N, GS-LSH project MIMIMI 2023-2025 for C.N Agence Nationale de la Recherche, MetaboInov, ANR-24-CE14-6636-01 for C.B ODYSSÉA association, 2020-2025 for C.N Association PROLIFIC, SYBU project 2019-2025 for C.N AIRC, Italy, IG-23670 for P.P Progetti di Rilevante Interesse Nazionale, PRIN20227Z2XRB to M.B, 2020RRJP5L, 202259LHXM, P2022WY85K_001, PNRR-CN00000041 to P.P

Abstract Background Triple-negative breast cancer (TNBC) is a clinically aggressive breast cancer subtype. It is a heterogeneous disease that remains difficult to stratify and that still lacks durable and biomarker-guided therapeutic options. Low expression of the tumour suppressor MTUS1 is associated with aggressive breast cancer features, but the biological properties of MTUS1 -low TNBC remain insufficiently defined. Our goal was to determine whether low MTUS1 expression defines shared proliferative and stress-adaptation mechanisms that could guide candidate therapeutic strategies and corresponding target/drug pairs in MTUS1 -low TNBC. Methods We labelled tumours from seven public TNBC RNA-seq cohorts based on the lowest and highest MTUS1 expression tertiles. Differential gene expression was analysed using gene set enrichment analysis (GSEA) on the Hallmark pathway database to identify deregulated biological pathways between MTUS1 -low TNBC tumours and their MTUS1 -high counterparts. Reproducibility was examined across independent TNBC cohorts and secondarily in broader breast cancer and selected TCGA tumour cohorts. Gene essentiality scores from CRISPR–Cas9 experiments in TNBC cell-line models were correlated to MTUS1 expression in these cell lines, to propose therapeutic strategies and their corresponding candidate target/drug pairs. Results MTUS1 -low tumours showed a reproducible pathway-level proliferation mechanism driven by the MYC oncogene and sustained by up-regulated oxidative phosphorylation, combined with stress adaptation mechanisms involving unfolded protein response (UPR), and DNA repair Hallmark gene sets. Based on CRISPR data, we propose 3 therapeutic strategies: (1) targeting MYC to reduce its transcriptional activity, (2) targeting proteins from UPR, (3) targeting DNA-repair. We also propose corresponding candidate target/drug pairs to allow experimental validation of these strategies. Conclusions Proliferation in low MTUS1 TNBC is driven by MYC and stress-adaptation mechanisms. By linking this tumour profile to CRISPR-derived dependency signals, our analysis prioritises experimentally testable target–pathway hypotheses centred on MYC, UPR/proteostasis, and DNA-repair or checkpoint control. Although the proposed therapeutic strategies and candidate targets remain to be experimentally tested, the latter finding is consistent with published work showing that ATIP3-deficient TNBC cell line models are sensitive to inhibition of the WEE1 PKMYT1 G2/M checkpoint kinases.

ATIP3-deficient breast cancers represent a subset of aggressive tumors with limited therapeutic options and poor prognosis. Here, we screened a panel of cell cycle kinase inhibitors to identify novel targets for these tumors. We show that loss of ATIP3 sensitizes breast cancer cells to WEE1 inhibition, resulting in aberrant mitoses characterized by detachment of centromere proteins from DNA and chromosome pulverization. This phenotype arises from excessive replication stress and DNA damage in S-phase, combined with premature mitotic entry driven by untimely CDK1 activation. Mechanistically, we identify DNA2 helicase/nuclease as a key mediator of chromosome pulverization. Importantly, the heightened sensitivity of ATIP3-deficient cells to WEE1 inhibition provides a strong rationale for clinical exploration of WEE1-targeted therapies. Furthermore, combining WEE1 and PKMYT1 inhibitors enhances therapeutic efficacy, offering a promising strategy for personalized treatment in ATIP3-deficient breast cancers.

Mitochondrial trafficking is reprogrammed in metastatic breast cancer cells to sustain their migratory and invasive behavior. Mitochondria repositioning to sites of high energy demand is governed by a balance between opposing dynein and kinesin-1 (KIF5B) molecular motors whose regulation remains incompletely understood. Here, we identify the SYBU gene as a candidate prognostic marker downregulated in metastatic disease. SYBU encodes syntabulin, a mitochondria outer membrane protein that interacts with dynein to counterbalance KIF5B-dependent anterograde transport to the cell cortex. Loss of SYBU disrupts the balance, causing excessive KIF5B-driven mitochondria movement, microtubule damage and deacetylation. In turn, microtubule deacetylation reinforces KIF5B-mediated transport, creating a positive feedback loop that drives mitochondria distribution close to the cell periphery and enhances cancer cell migration. Pharmacological inhibition of the tubulin deacetylase HDAC6 restores mitochondrial positioning and reduces cell migration in SYBU -deficient cells. Our findings identify SYBU as a key regulator of mitochondrial trafficking and pave the way to personalized therapeutic approaches for metastatic breast tumors with low SYBU expression. ### Competing Interest Statement The authors have declared no competing interest. France Bio imaging (FBI), ANR-24-INBS-0005 FBI BIOGEN La Ligue Contre le Cancer, https://ror.org/00rkrv905, MM PhD thesis (2021-2025) Entreprises contre le Cancer Paris GEFLUC, SYBU project (2024-2025) Fondation ARC pour la Recherche sur le Cancer, https://ror.org/0489qz649, HM post-doc (2017-2021) Fondation de Rothschild, https://ror.org/02yfw7119, SYBU project (2023-2025) AG2R LA MONDIALE, 2019-2021 the Emergence call from the Labex LERMIT of Paris-Saclay University, Project Mito-Micro Odyssea, MT and breast cancer Prolific, MT-BK Ruban Rose, Nahmias 2022

ATIP3, a microtubule-associated protein with tumor suppressor functions, is frequently lost in aggressive breast cancers, particularly in 60-80% of triple-negative breast cancer (TNBC), where its absence correlates with poor prognosis. ATIP3 deficiency is associated with heightened aneuploidy, presenting a vulnerability exploitable by targeted therapies. WEE1 kinase, a critical regulator of the G2/M checkpoint, has demonstrated therapeutic promise in this context. Inhibiting WEE1 with AZD1775 selectively compromises ATIP3-deficient cells by exacerbating DNA damage, mitotic abnormalities and chromosome pulverization, ultimately leading to cell death. However, the clinical application of WEE1 inhibitors is constrained by dose-dependent toxicities, necessitating combination strategies to enhance efficacy while reducing toxicity. To address these challenges, we explored the complementary targeting of PKMYT1, another G2/M checkpoint kinase with distinct yet overlapping functions to WEE1. Using 3D cultures of breast cancer cell lines, we compared the responses of ATIP3-deficient and ATIP3-expressing cells to three PKMYT1 inhibitors—RP-6306, GSK1520489A, and MYT1-IN-I. ATIP3-deficient cells exhibited heightened sensitivity to all three inhibitors, particularly RP-6306, which achieved significantly lower IC50 values. To maximize therapeutic impact, we combined RP-6306 with AZD1775, resulting in a synergistic reduction in cell viability in ATIP3-deficient cells. Dual inhibition of WEE1 and PKMYT1 intensified replication stress, DNA damage, and abnormal mitoses, exceeding the effects observed with either inhibitor alone. Importantly, the dual-targeting approach enables the use of lower doses of WEE1 inhibitors, potentially mitigating the toxicities associated with high-dose monotherapies. Our findings establish PKMYT1 as a complementary target to WEE1 in ATIP3-deficient breast cancers. The synergistic effects of their combined inhibition not only enhance therapeutic efficacy but also address key limitations of current WEE1-based therapies. This dual-targeting strategy holds promise for improving outcomes in aggressive breast cancer subtypes. Furthermore, our work highlights the potential of ATIP3 as a predictive biomarker for patient stratification and the rational development of personalized therapies. Maria M. Haykal, Marie Varin de Lapeyriere, Alexis Denis, Sylvie Rodrigues-Ferreira, Clara Nahmias. Dual inhibition of PKMYT1 and WEE1 kinases as a targeted therapy for ATIP3-deficient breast cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6964.

A wide panel of microtubule-associated proteins and kinases is involved in coordinated regulation of the microtubule cytoskeleton and may thus represent valuable molecular markers contributing to major cellular pathways deregulated in cancer. We previously identified a panel of 17 microtubule-related (MT-Rel) genes that are differentially expressed in breast tumors showing resistance to taxane-based chemotherapy. In the present study, we evaluated the expression, prognostic value and functional impact of these genes in breast cancer. We show that 14 MT-Rel genes (KIF4A, ASPM, KIF20A, KIF14, TPX2, KIF18B, KIFC1, AURKB, KIF2C, GTSE1, KIF15, KIF11, RACGAP1, STMN1) are up-regulated in breast tumors compared with adjacent normal tissue. Six of them (KIF4A, ASPM, KIF20A, KIF14, TPX2, KIF18B) are overexpressed by more than 10-fold in tumor samples and four of them (KIF11, AURKB, TPX2 and KIFC1) are essential for cell survival. Overexpression of all 14 genes, and underexpression of 3 other MT-Rel genes (MAST4, MAPT and MTUS1) are associated with poor breast cancer patient survival. A Systems Biology approach highlighted three major functional networks connecting the 17 MT-Rel genes and their partners, which are centered on spindle assembly, chromosome segregation and cytokinesis. Our studies identified mitotic Aurora kinases and their substrates as major targets for therapeutic approaches against breast cancer.

Supplementary Information PDF file - 115K, Supplemental materials and methods, legends to Movies and supplemental references

Supplementary Tables XLS file - 200K, Table SI : breast tumors and normal samples data; Table SII: MTUS1 expression level in invasive breast tumors collected at Institut Gustave Roussy; Table SIII: Time-course of metastasis formation

Movie 6 AVI file - 5618K, Time-lapse videomicroscopy of EB3-GFP comets in wild type and mCherry-ATIP3 transfected MRC5 cells

Aneuploidy, a hallmark of cancer, is a prominent feature associated with poor prognosis in breast cancer. Here, we screened a panel of cell cycle kinase inhibitors to identify novel targets for highly aneuploid breast cancers. We show that increasing aneuploidy in breast cancer cells sensitizes to the inhibition of WEE1 kinase. Upon exposure to WEE1 inhibitor, aneuploid cells exhibit aberrant mitosis characterized by the detachment of centromere proteins from centromeric DNA and pulverization of chromosomes. The occurrence of such phenotype is driven by excessive levels of replication stress and DNA damage during S-phase, that in turn trigger major defects in the subsequent mitosis. We show that DNA2 helicase/nuclease, that regulates replication of centromeric DNA, is the key player responsible for severe chromosome pulverization in mitosis. The heightened vulnerability of aneuploid cells to WEE1 inhibition, coupled with underlying molecular mechanisms, provides a rationale for clinical exploration of WEE1-targeted therapies against aneuploid breast cancers. Impact Statement Increased vulnerability of aneuploid cells to WEE1 inhibition is orchestrated by the DNA2 nuclease/helicase. These findings open new therapeutic strategies in the context of personalized medicine in breast cancer.

Supplementary Figures PDF file - 486K, Figure S1. Low levels of ATIP3 associate with decreased overall and relapse-free survival of the patients;Figure S2. Anti-metastatic effects of ATIP3 in vivo; Figure S3. ATIP3 inhibits breast cancer cell proliferation and migration; Figure S4. ATIP3 regulates MT dynamics; Figure S5. Characterization of ATIP3 domains

Breast cancer is one of the most frequent malignancies among women worldwide. Based on clinical and molecular features of breast tumors, patients are treated with chemotherapy, hormonal therapy and/or radiotherapy and more recently with immunotherapy or targeted therapy. These different therapeutic options have markedly improved patient outcomes. However, further improvement is needed to fight against resistance to treatment. In the rapidly growing area of research for personalized medicine, predictive biomarkers - which predict patient response to therapy - are essential tools to select the patients who are most likely to benefit from the treatment, with the aim to give the right therapy to the right patient and avoid unnecessary overtreatment. The search for predictive biomarkers is an active field of research that includes genomic, proteomic and/or machine learning approaches. In this review, we describe current strategies and innovative tools to identify, evaluate and validate new biomarkers. We also summarize current predictive biomarkers in breast cancer and discuss companion biomarkers of targeted therapy in the context of precision medicine.

La reprogrammation métabolique est l’un des marqueurs de la carcinogenèse. Au cœur de cette reprogrammation se trouvent les mitochondries qui produisent l’énergie sous forme de molécules d’ATP. La régulation spatio-temporelle de la production d’ATP, indispensable pour fournir l’énergie au bon endroit et au bon moment, est assurée par le transport intracellulaire des mitochondries. Les complexes Miro/TRAK présents à la surface des mitochondries se lient aux protéines motrices de la cellule (dynéine, kinésine, myosine) pour transporter les mitochondries le long du cytosquelette. Ces acteurs du transport mitochondrial sont souvent dérégulés dans le cancer. Nous présentons dans cette revue les mécanismes par lesquels le transport mitochondrial contribue à la migration, à la division cellulaire et à la réponse au stress des cellules cancéreuses. Décrypter ces mécanismes pourrait ouvrir la voie à de nouvelles approches thérapeutiques en oncologie.

Cancer cells are characterized by a deregulation of their metabolic activity, which allows them to meet a high energy demand. Mitochondria are key organelles that control several metabolic processes and represent the main source of energy in the form of ATP. Intracellular transport of mitochondria is essential for addressing these organelles to the right place at the right time according to energy requirement. Mitochondrial transport in cancer cells involves mitochondria-associated Miro/TRAK complexes that bind to motor proteins (kinesins, dyneins and myosins) to promote mitochondrial displacement along microtubules or actin filaments. This review focuses on the molecular players of intracellular mitochondrial transport along microtubules during cell migration and mitosis, and their deregulation in tissues from cancer patients. Intercellular mitochondrial transport upon cancer cell exposure to hypoxia or chemotherapy is also presented. This field of investigation opens new interesting perspectives in oncology, as targeting mitochondrial transport may represent an innovative strategy for treating cancer.

Breast cancer is the leading cause of death by malignancy among women worldwide. Clinical data and molecular characteristics of breast tumors are essential to guide clinician’s therapeutic decisions. In the new era of precision medicine, that aims at personalizing the treatment for each patient, there is urgent need to identify robust companion biomarkers for new targeted therapies. This review focuses on ATIP3, a potent anti-cancer protein encoded by candidate tumor suppressor gene MTUS1, whose expression levels are markedly down-regulated in breast cancer. ATIP3 is a microtubule-associated protein identified both as a prognostic biomarker of patient survival and a predictive biomarker of breast tumors response to taxane-based chemotherapy. We present here recent studies pointing out ATIP3 as an emerging anti-cancer protein and a potential companion biomarker to be combined with future personalized therapy against ATIP3-deficient breast cancer.

Carcinogenesis is a multi-step process that refers to transformation of a normal cell into a tumoral neoplastic cell. The mechanisms that promote tumor initiation, promotion and progression are varied, complex and remain to be understood. Studies have highlighted the involvement of oncogenic mutations, genomic instability and epigenetic alterations as well as metabolic reprogramming, in different processes of oncogenesis. However, the underlying mechanisms still have to be clarified. Mitochondria are central organelles at the crossroad of various energetic metabolisms. In addition to their pivotal roles in bioenergetic metabolism, they control redox homeostasis, biosynthesis of macromolecules and apoptotic signals, all of which are linked to carcinogenesis. In the present review, we discuss how mitochondria contribute to the initiation of carcinogenesis through gene mutations and production of oncometabolites, and how they promote tumor progression through the control of metabolic reprogramming and mitochondrial dynamics. Finally, we present mitochondrial metabolism as a promising target for the development of novel therapeutic strategies.

Taxanes are microtubule-targeting drugs used as cytotoxic chemotherapy to treat most solid tumors. The development of resistance to taxanes is a major cause of therapeutic failure and overcoming chemoresistance remains an important challenge to improve patient's outcome. Extensive efforts have been made recently to identify predictive biomarkers to select populations of patients who will benefit from taxane-based chemotherapy and avoid inefficient treatment of patients with innate resistance. This, together with the discovery of new mechanisms of resistance that include metabolic reprogramming and dialogue between tumor and its microenvironment, pave the way to a new era of personalized medicine. In this review, we recapitulate recent insights into taxane resistance and present promising emerging strategies to overcome chemoresistance in the future.

Aneuploidy, an abnormal chromosome number, is a hallmark of cancer. We recently showed that depletion of microtubule-associated protein ATIP3 (AT2 receptor-interacting protein 3) induces aneuploidy and sensitizes breast cancer cells to taxanes. Combining taxane treatment with ATIP3 depletion cooperates to reach a detrimental level of aneuploidy.

Taxane-based chemotherapy is frequently used in neoadjuvant treatment of breast cancer patients to reduce tumor growth and lymph node metastasis. However, few patients benefit from chemotherapy and predictive biomarkers of chemoresistance are needed. The microtubule-associated protein ATIP3 has recently been identified as a predictive biomarker whose low levels in breast tumors are associated with increased sensitivity to chemotherapy. In this study, we investigated whether ATIP3 deficiency may impact the effects of paclitaxel on cancer cell migration and lymph node metastasis. Expression levels of ATIP3 were analyzed in a cohort of 133 breast cancer patients and classified according to lymph node positivity following neoadjuvant chemotherapy. Results showed that low ATIP3 levels are associated with reduced axillary lymph node metastasis. At the functional level, ATIP3 depletion increases cell migration, front-rear polarity and microtubule dynamics at the plus ends, but paradoxically sensitizes cancer cells to the inhibitory effects of paclitaxel on these processes. ATIP3 silencing concomitantly increases the incorporation of fluorescent derivative of Taxol along the microtubule lattice. Together our results support a model in which alterations of microtubule plus ends dynamics in ATIP3-deficient cells may favor intracellular accumulation of paclitaxel, thereby accounting for increased breast tumor sensitivity to chemotherapy.

Cancer is a complex disease and it is now clear that not only epithelial tumor cells play a role in carcinogenesis. The tumor microenvironment is composed of non-stromal cells, including endothelial cells, adipocytes, immune and nerve cells, and a stromal compartment composed of extracellular matrix, cancer-associated fibroblasts and mesenchymal cells. Tumorigenesis is a dynamic process with constant interactions occurring between the tumor cells and their surroundings. Even though all connections have not yet been discovered, it is now known that crosstalk between actors of the microenvironment drives cancer progression. Taking into account this complexity, it is important to develop relevant models to study carcinogenesis. Conventional 2D culture models fail to represent the entire tumor microenvironment properly and the use of animal models should be decreased with respect to the 3Rs rule. To this aim, in vitro organotypic models have been significantly developed these past few years. These models have different levels of complexity and allow the study of tumor cells alone or in interaction with the microenvironment actors during the multiple stages of carcinogenesis. This review depicts recent insights into organotypic modeling of the tumor and its microenvironment all throughout cancer progression. It offers an overview of the crosstalk between epithelial cancer cells and their microenvironment during the different phases of carcinogenesis, from the early cell autonomous events to the late metastatic stages. The advantages of 3D over classical 2D or in vivo models are presented as well as the most promising organotypic models. A particular focus is made on organotypic models used for studying cancer progression, from the less complex spheroids to the more sophisticated body-on-a-chip. Last but not least, we address the potential benefits of these models in personalized medicine which is undoubtedly a domain paving the path to new hopes in terms of cancer care and cure.