High tumour-infiltrating lymphocytes correlate with distinct gene expression profile and favourable survival in single hormone receptor-positive breast cancer.

Introduction: This study aimed to evaluate the impact of tumour-infiltrating lymphocytes (TILs) on the expression of immune-related genes and prognosis in single hormone receptor-positive breast cancer. Material and methods: Tumour-infiltrating lymphocytes were analysed according to the guidelines of the International TILs Working Group in a cohort of 206 patients with single hormone receptor-positive breast cancer. Of these, 44.7% were classified as ER+/PgR-/HER2-, 18.4% as ER+/PgR-/HER2+, 26.2% as ER-/PgR+/HER2-, and 10.7% as ER-/PgR+/HER2+. Moreover, in 52 samples the analysis of gene expression profiling was performed using nCounter technology. Results: Most cases (74.3%) showed at least 1% of stromal TILs, with a median of 4%, mean of 16.3%, and interquartile range of 0-20%. ER-/PgR+ tumours displayed significantly higher TILs density than ER+/PgR- cases (p < 0.001, Wilcoxon test), regardless of HER2 status. The abundance of TILs was positively associated with ER-/PgR+ phenotype, higher Ki-67, and higher grade, but not with age, tumour size, or regional and distant metastases at diagnosis. Additionally, in ER+/PgR- subgroup higher TILs were associated with HER2-positive status. Stromal TILs > 5% were associated with better survival in the whole group, but this effect was less prominent in ER-/PgR+ patients. We identified 50 differentially expressed genes (DEGs) between single hormone receptor-positive breast tumours with high and low TILs, including 39 up-regulated and 11 down-regulated genes in the high TILs group. Conclusions: The up-regulated expression of immune-related genes was consistent also among separately analysed single hormone receptor-positive groups (ER+/PgR- and ER-/PgR+).

Chasing Graphene-Based Anticancer Drugs: Where are We Now on the Biomedical Graphene Roadmap?

Graphene and graphene-based materials have attracted growing interest for potential applications in medicine because of their good biocompatibility, cargo capability and possible surface functionalizations. In parallel, prototypic graphene-based devices have been developed to diagnose, imaging and track tumor growth in cancer patients. There is a growing number of reports on the use of graphene and its functionalized derivatives in the design of innovative drugs delivery systems, photothermal and photodynamic cancer therapy, and as a platform to combine multiple therapies. The aim of this review is to introduce the latest scientific achievements in the field of innovative composite graphene materials as potentially applied in cancer therapy. The "Technology and Innovation Roadmap" published in the Graphene Flagship indicates, that the first anti-cancer drugs using graphene and graphene-derived materials will have appeared on the market by 2030. However, it is necessary to broaden understanding of graphene-based material interactions with cellular metabolism and signaling at the functional level, as well as toxicity. The main aspects of further research should elucidate how treatment methods (e.g., photothermal therapy, photodynamic therapy, combination therapy) and the physicochemical properties of graphene materials influence their ability to modulate autophagy and kill cancer cells. Interestingly, recent scientific reports also prove that graphene nanocomposites modulate cancer cell death by inducing precise autophagy dysfunctions caused by lysosome damage. It turns out as well that developing photothermal oncological treatments, it should be taken into account that near-infrared-II radiation (1000-1500 nm) is a better option than NIR-I (750-1000 nm) because it can penetrate deeper into tissues due to less scattering at longer wavelengths radiation.

Insights into Kidney Dysplasia in Duplex Kidneys: From Radiologic Diagnosis to Histopathologic Understanding.

Duplex kidney is a urinary tract anomaly commonly associated with a wide range of primary and secondary parenchymal structural abnormalities. We present a unique comparison of US and MRI findings with histopathology following partial resection of duplex kidneys due to nephropathy. We examined a group of 21 children with duplex kidneys who were qualified for heminephrectomy (24 kidney units (KU)). All patients underwent US and MRI prior to the surgery. The imaging results were compared with histopathologic findings. In 21/24 KU, dysplastic changes were found on histopathology, including all with obstructive nephropathy and 7/10 specimens with refluxing uropathy. The loss of corticomedullary differentiation on US and increased signal on T2-weighted images (T2WI) on MRI were the imaging findings that best correlated with fibrosis. In children with megaureter, there were no statistical differences in histopathological findings between primary megaureter, megaureter with ureterocele, and megaureter with ectopia (p > 0.05). The extent of dysplasia of the affected pole correlated negatively with residual function in MRI. Kidney dysplasia and inflammation in the kidney with obstructive nephropathy are the most important histopathologic findings of this study. US is a valuable screening tool, and MRI enables morphologic and functional assessments of the nephropathy in duplex kidneys.

Recommendations of the Polish Association of Neuropathologists on performing post-mortem examination of the brain and spinal cord.

Neuropathological central nervous system (CNS) post-mortem examination is a highly specialistic element of the autopsy procedure with methodological specificity. Herein we propose updated recommendations for CNS autopsy for pathologists and neuropathologists. The protocol includes the compendium of neuroanatomy with current nomenclature, consecutive steps of gross examination, as well as appropriate sampling algorithms in different clinical and pathological settings. The significance of pathoclinical cooperation in differential diagnosis is exposed. We believe it is essential to create and promote the guidelines to improve the quality of CNS post-mortem examination at the national level.

Targeted sequencing of cancer-related genes reveals a recurrent TOP2A variant which affects DNA binding and coincides with global transcriptional changes in glioblastoma.

High-grade gliomas are aggressive, deadly primary brain tumors. Median survival of patients with glioblastoma (GBM, WHO grade 4) is 14 months and <10% of patients survive 2 years. Despite improved surgical strategies and forceful radiotherapy and chemotherapy, the prognosis of GBM patients is poor and did not improve over decades. We performed targeted next-generation sequencing with a custom panel of 664 cancer- and epigenetics-related genes, and searched for somatic and germline variants in 180 gliomas of different WHO grades. Herein, we focus on 135 GBM IDH-wild type samples. In parallel, mRNA sequencing was accomplished to detect transcriptomic abnormalities. We present the genomic alterations in high-grade gliomas and the associated transcriptomic patterns. Computational analyses and biochemical assays showed the influence of TOP2A variants on enzyme activities. In 4/135 IDH-wild type GBMs we found a novel, recurrent mutation in the TOP2A gene encoding topoisomerase 2A (allele frequency [AF] = 0.03, 4/135 samples). Biochemical assays with recombinant, wild type (WT) and variant proteins demonstrated stronger DNA binding and relaxation activity of the variant protein. GBM patients carrying the altered TOP2A had shorter overall survival (median OS 150 vs 500 days, P = .0018). In the GBMs with the TOP2A variant we found transcriptomic alterations consistent with splicing dysregulation. luA novel, recurrent TOP2A mutation, which was found exclusively in four GBMs, results in the TOP2A E948Q variant with altered DNA binding and relaxation activities. The deleterious TOP2A mutation resulting in transcription deregulation in GBMs may contribute to disease pathology.

Analysis of BRCA1 and BRCA2 alternative splicing in predisposition to ovarian cancer.

BACKGROUND: The mRNA splicing is regulated on multiple levels, resulting in the proper distribution of genes' transcripts in each cell and maintaining cell homeostasis. At the same time, the expression of alternative transcripts can change in response to underlying genetic variants, often missed during routine diagnostics. AIM: The main aim of this study was to define the frequency of aberrant splicing in BRCA1 and BRCA2 genes in blood RNA extracted from ovarian cancer patients who were previously found negative for the presence of pathogenic alterations in the 25 most commonly analysed ovarian cancer genes, including BRCA1 and BRCA2. MATERIAL AND METHODS: Frequency and spectrum of splicing alterations in BRCA1 and BRCA2 genes were analysed in blood RNA from 101 ovarian cancer patients and healthy controls (80 healthy women) using PCR followed by gel electrophoresis and Sanger sequencing. The expression of splicing events was examined using RT-qPCR. RESULTS: We did not identify any novel, potentially pathogenic splicing alterations. Nevertheless, we detected six naturally occurring transcripts, named BRCA1ΔE9-10, BRCA1ΔE11, BRCA1ΔE11q, and BRCA2ΔE3, BRCA2ΔE12 and BRCA2ΔE17-18 of which three (BRCA1ΔE11q, BRCA1ΔE11 and BRCA2ΔE3) were significantly higher expressed in the ovarian cancer cohort than in healthy controls (p ≤ 0.0001). CONCLUSIONS: This observation indicates that the upregulation of selected naturally occurring transcripts can be stimulated by non-genetic mechanisms and be a potential systemic response to disease progression and/or treatment. However, this hypothesis requires further examination.

Neuropeptide Y and its receptors in prostate cancer: associations with cancer invasiveness and perineural spread.

PURPOSE: Neuropeptide Y (NPY) is a pleiotropic peptide, which is involved in many biological mechanisms important in regulation of cell growth and survival. The aim of this study was a comprehensive analysis of the NPY system in prostate pathology. METHODS: The study was based on immunohistochemical analysis of NPY and its receptors, Y1R, Y2R and Y5R, in tissue samples from benign prostate (BP), primary prostate cancer (PCa) and PCa bone metastases. Tissue microarray (TMA) technique was employed, with analysis of multiple cores from each specimen. Intensity of the immunoreactivity and expression index (EI), as well as distribution of the immunostaining in neoplastic cells and stromal elements were evaluated. Perineural invasion (PNI) and extraprostatic extension (EPE) were areas of special interests. Moreover, a transwell migration assay on the LNCaP PCa cell line was used to assess the chemotactic properties of NPY. RESULTS: Morphological analysis revealed homogeneous membrane and cytoplasmic pattern of NPY staining in cancer cells and its membrane localization with apical accentuation in BP glands. All elements of the NPY system were upregulated in pre-invasive prostate intraepithelial neoplasia, PCa and metastases. EI and staining intensity of NPY receptors were significantly higher in PCa then in BP with correlation between Y2R and Y5R. The strength of expression of the NPY system was further increased in the PNI and EPE areas. In bone metastases, Y1R and Y5R presented high expression scores. CONCLUSION: The results of our study suggest that the NPY system is involved in PCa, starting from early stages of its development to disseminated states of the disease, and participates in the invasion of PCa into the auto and paracrine matter.

Parenchyma Sparing Anatomic Liver Resections (Bi- and Uni-Segmentectomies) for Liver Tumours in Children-A Single-Centre Experience.

Purpose: To present a single-centre experience in bi- and uni-segmentectomies for primary liver tumours in children. METHODS: This study included 23 patients that underwent (bi)segmentectomy. There were 15 malignant tumours (hepatoblastoma-13 patients), 7 benign tumours, and 1 calcifying nested stromal epithelial tumour. RESULTS: The median tumour diameter was 52 mm (range 15-170 mm). Bisegmentectomy 2-3 was most frequently performed (seven patients), followed by bisegmentectomy 5-6 (four patients). The median operative time was 225 min (range 95-643 min). Intraoperative complications occurred in two patients-small bowel perforation in one and an injury of the small peripheral bile duct resulting in biloma in the other. The median resection margin in patients with hepatoblastoma was 3 mm (range 1-15 mm). Microscopically negative margin status was achieved in 12 out of 13 patients. There were two recurrences. After a median follow-up time of 38 months (range 12-144 months), all 13 patients with HB were alive with no evidence of disease. Two relapsed patients were alive with no evidence of disease. CONCLUSIONS: From the available literature and data presented here, we propose that (bi)segmentectomy can become a viable surgical option in carefully selected paediatric patients and is sufficient to achieve a cure. Further studies evaluating the impact of parenchymal preservation surgery on surgical and oncological outcome should be conducted with a larger dataset.

"Neuropatologia Polska". The journal and its topics in the first decade of existence (1963-1972).

In this article authors would like to present the history of the "Neuropatologia Polska" journal (since 1994: "Folia Neuropathologica") in its first decade of existence. It outlines the circumstances surrounding the creation of the journal and shows how it evolved in the first years. The vast material analysed from the consecutive issues of the journal in the years from 1963 to 1972 was subjected to statistical and content analysis. From its first year, the journal has included works of a very high substantive level and a wide range of topics. The authors presented the results of contemporary research in many areas. The "Neuropatologia Polska" journal (later "Folia Neuropathologica") set paths for the development of neuropathology in clinical and experimental aspects. What is very important, it created a platform for international cooperation in many fields, included researchers and scientists from Western countries and foreign academic centres in difficult times. This article was created on the 60th anniversary of creation of "Neuropatologia Polska".

Pathway-level mutation analysis in primary high-grade serous ovarian cancer and matched brain metastases.

Brain metastases (BMs) in ovarian cancer (OC) are a rare event. BMs occur most frequently in high-grade serous (HGS) OC. The molecular features of BMs in HGSOC are poorly understood. We performed a whole-exome sequencing analysis of ten matched pairs of formalin-fixed paraffin-embedded samples from primary HGSOC and corresponding BMs. Enrichment significance (p value; false discovery rate) was computed using the Reactome, the Kyoto Encyclopedia of Genes and Genomes pathway collections, and the Gene Ontology Biological Processes. Germline DNA damage repair variants were found in seven cases (70%) and involved the BRCA1, BRCA2, ATM, RAD50, ERCC4, RPA1, MLHI, and ATR genes. Somatic mutations of TP53 were found in nine cases (90%) and were the only stable mutations between the primary tumor and BMs. Disturbed pathways in BMs versus primary HGSOC constituted a complex network and included the cell cycle, the degradation of the extracellular matrix, cell junction organization, nucleotide metabolism, lipid metabolism, the immune system, G-protein-coupled receptors, intracellular vesicular transport, and reaction to chemical stimuli (Golgi vesicle transport and olfactory signaling). Pathway analysis approaches allow for a more intuitive interpretation of the data as compared to considering single-gene aberrations and provide an opportunity to identify clinically informative alterations in HGSOC BM.

Alterations in key signaling pathways in sinonasal tract melanoma. A molecular genetics and immunohistochemical study of 90 cases and comprehensive review of the literature.

Sinonasal mucosal melanoma is a rare tumor arising within the nasal cavity, paranasal sinuses, or nasopharynx (sinonasal tract). This study evaluated 90 cases diagnosed in 29 males and 61 females with median age 68 years. Most tumors involved the nasal cavity and had an epithelioid morphology. Spectrum of research techniques used in this analysis includes targeted-DNA and -RNA next-generation sequencing, Sanger sequencing, fluorescence in situ hybridization and immunohistochemistry. Sinonasal melanomas were commonly driven by RAS (38/90, 42%), especially NRAS (n = 36) mutations and rarely (4/90, 4%) displayed BRAF pathogenic variants. BRAF/RAS mutants were more frequent among paranasal sinuses (10/14, 71%) than nasal (26/64, 41%) tumors. BRAF/RAS-wild type tumors occasionally harbored alterations of the key components and regulators of Ras-MAPK signaling pathway: NF1 mutations (1/17, 6%) or NF1 locus deletions (1/25, 4%), SPRED1 (3/25, 12%), PIK3CA (3/50, 6%), PTEN (4/50, 8%) and mTOR (1/50, 2%) mutations. These mutations often occurred in a mutually exclusive manner. In several tumors some of which were NRAS mutants, TP53 was deleted (6/48, 13%) and/or mutated (5/90, 6%). Variable nuclear accumulation of TP53, mirrored by elevated nuclear MDM2 expression was seen in >50% of cases. Furthermore, sinonasal melanomas (n = 7) including RAS/BRAF-wild type tumors (n = 5) harbored alterations of the key components and regulators of canonical WNT-pathway: APC (4/90, 4%), CTNNB1 (3/90, 3%) and AMER1 (1/90, 1%). Both, TERT promoter mutations (5/53, 9%) and fusions (2/40, 5%) were identified. The latter occurred in BRAF/RAS-wild type tumors. No oncogenic fusion gene transcripts previously reported in cutaneous melanomas were detected. Eight tumors including 7 BRAF/RAS-wild type cases expressed ADCK4::NUMBL cis-fusion transcripts. In summary, this study documented mutational activation of NRAS and other key components and regulators of Ras-MAPK signaling pathway such as SPRED1 in a majority of sinonasal melanomas.

WHO CNS5 2021 classification of gliomas: a practical review and road signs for diagnosing pathologists and proper patho-clinical and neuro-oncological cooperation.

The 5th edition of World Health Organization (WHO) Central Nervous System (CNS) tumours classification has transformed the pathological diagnosis of gliomas from purely histological to the multilayered integrated one with molecular biomarkers necessary for proper classification, risk stratification, and prognostic-predictive clinical purposes. Because of deep and important changes in taxonomy and diagnostic approach to gliomas, this manuscript is a review of WHO CNS classification 5th edition with general testing guidance for pathologists and clinicians working in neuro-oncology.

Antibody-Drug Conjugates in Uro-Oncology.

Currently available treatment options for patients with refractory metastatic prostate, bladder, or kidney cancers are limited with the prognosis remaining poor. Advances in the pathobiology of tumors has led to the discovery of cancer antigens that may be used as the target for cancer treatment. Antibody-drug conjugates (ADCs) are a relatively new concept in cancer treatment that broaden therapeutic landscape. ADCs are examples of a 'drug delivery into the tumor' system composed of an antigen-directed antibody linked to a cytotoxic drug that may release cytotoxic components after binding to the antigen located on the surface of tumor cells. The clinical properties of drugs are influenced by every component of ADCs. Regarding uro-oncology, enfortumab vedotin (EV) and sacituzumab govitecan (SG) are currently registered for patients with locally advanced or metastatic urothelial cancer following previous treatment with an immune checkpoint inhibitor (iCPI; programmed death receptor-1 [PD-1] or programmed death-ligand 1 [PD-L1]) inhibitor) and platinum-containing chemotherapy. The EV-301 trial showed that EV significantly prolonged the overall survival compared with classic chemotherapy. The TROPHY-U-01 trial conducted to evaluate SG demonstrated promising results as regards the objective response rate and duration of response. The safety and efficacy of ADCs in monotherapy and polytherapy (mainly with iCPIs) for different cancer stages and tumor types are assessed in numerous ongoing clinical trials. The aim of this review is to present new molecular biomarkers, specific mechanisms of action, and ongoing clinical trials of ADCs in genitourinary cancers. In the expert discussion, we assess the place of ADCs in uro-oncology and discuss their clinical value.

Hypoxic hepatitis as a complication of newly diagnosed type 1 diabetes in a teenager.

Hypoxic hepatitis is a rare complication of type 1 diabetes with unknown prevalence in Pediatrics. We present a case report of an 11-year-old boy admitted to the ER in the spring of 2020 (the beginning of the COVID19 pandemic in Poland) due to nausea, abdominal pain, and weight loss. A diagnosis of type 1 diabetes accompanied by severe ketoacidosis (pH 6.9, blood glucose 632mg/dl, ketone bodies in urine - 150mg/dl) was made. The hyperglycemia, ketoacidosis, and water-electrolyte disturbances were treated in the Pediatric Intensive Care Unit. On day 4, the boy developed fulminant septic shock with high aminotransferases (AST 9026 U/l, ALT 3559 U/l). CT scan revealed hepatic enlargement and steatosis. Acute viral hepatitis was suspected. The levels of anti-CMV IgM and IgG antibodies were slightly elevated. At autopsy, the liver was enlarged, with petechial bleedings on the surface. The liver parenchyma was congested, with signs of steatosis. Microscopically, there was extensive centrilobular necrosis, acute passive sinusoidal congestion, and steatosis of hepatocytes. There were no signs of CMV infection. Based on the entire clinicopathological picture, the patient was diagnosed with hypoxic hepatitis, complicated by septic shock and multiple organ failure.

Hypoxia-activated neuropeptide Y/Y5 receptor/RhoA pathway triggers chromosomal instability and bone metastasis in Ewing sarcoma.

Adverse prognosis in Ewing sarcoma (ES) is associated with the presence of metastases, particularly in bone, tumor hypoxia and chromosomal instability (CIN). Yet, a mechanistic link between these factors remains unknown. We demonstrate that in ES, tumor hypoxia selectively exacerbates bone metastasis. This process is triggered by hypoxia-induced stimulation of the neuropeptide Y (NPY)/Y5 receptor (Y5R) pathway, which leads to RhoA over-activation and cytokinesis failure. These mitotic defects result in the formation of polyploid ES cells, the progeny of which exhibit high CIN, an ability to invade and colonize bone, and a resistance to chemotherapy. Blocking Y5R in hypoxic ES tumors prevents polyploidization and bone metastasis. Our findings provide evidence for the role of the hypoxia-inducible NPY/Y5R/RhoA axis in promoting genomic changes and subsequent osseous dissemination in ES, and suggest that targeting this pathway may prevent CIN and disease progression in ES and other cancers rich in NPY and Y5R.

High expression of progesterone receptor may be an adverse prognostic factor in oestrogen receptor-negative/progesterone receptor-positive breast cancer: results of comprehensive re-evaluation of multi-institutional case series.

Oestrogen receptor (ER)-negative (-) progesterone receptor (PgR)-positive (+) is the least common combination of steroid receptor expression observed in breast cancer. There are many controversies regarding the actual existence of ER-/PgR+ phenotype. In the current study, we aimed to perform comprehensive immunohistochemical re-evaluation of ER-/PgR+ breast cancers from multiple institutions. A total of 135 cases of ER-/PgR+ breast cancer were collected from 11 institutions from the period 2006-2020 and subsequently stained with three clinically validated anti-ER antibody clones: SP1 (Roche), 1D5 (Dako), and EP1 (Dako), and two anti-PgR antibody clones: 636 (Dako), and 1E2 (Roche). Clinicopathological characteristics of confirmed and re-categorised cases were analysed. Seventy-six cases retained the original ER-/PgR+ phenotype, including 21 HER2+ and 55 HER2- tumours. Forty-seven cases were ER+ with at least one anti-ER antibody, and 12 cases were re-categorised as double-negatives across all anti-ER and anti-PgR antibodies. No significant differences in survival were observed between groups in the HER2+ category. In the HER2- cohort, confirmed ER-/PgR+, ER+ tumours with discrepant ER staining, and triple negatives had inferior overall survival compared to concordant ER+ cases. Progesterone receptor expression in >20% of cells was identified as an adverse prognostic factor in ER-/PgR+/HER2- breast cancer in a multivariable model adjusted by stage (HR 5.0, 95% CI 1.3-19.2, p=0.019). We performed one of the largest validation studies so far on ER-/PgR+ breast cancer and confirmed the existence of this subgroup. Moreover, we identified high PgR expression as an adverse prognostic factor.

Bilateral Ovarian Germ Cell Tumor in a 46,XX Female with Nijmegen Breakage Syndrome and Hypergonadotropic Hypogonadism

Nijmegen breakage syndrome (NBS) is a rare autosomal recessive disease, affecting mainly patients of Slavic origin. It is caused by a defect in the NBN gene, resulting in defective nibrin protein formation. This leads to chromosomal instability, which predisposes to cancer, with lymphoid malignancies predominating. Nibrin is also involved in gonadal development and its disfunction in females with NBS frequently results in a pure gonadal dysgenesis (PGD) causing hypergonadotropic hypogonadism. However, only a few ovarian tumors in NBS patients have been reported to date. We describe the first case of a girl with NBS with PGD, who developed metachronous bilateral ovarian germ cell tumors (dysgerminoma and gonadoblastoma). Pathogenesis of PGD, neoplastic transformation and therapeutic approach in females with NBS are discussed.

Central nervous system autopsy - a neuropathological procedure based on multidisciplinary pathoclinical cooperation.

INTRODUCTION: Neuropathological brain and spinal cord post mortem examination is a distinct procedure that still plays an important role in modern medicine. In front of increasing amounts of clinical and genetic data, together with important developments in the field of neuroimaging, the Polish Association of Neuropathologists have updated their recommendations regarding central nervous system (CNS) examination. These guidelines are aimed at neuropathologists, pathologists and clinicians. AIM OF THE STUDY: Presentation of the outlined recommendations as their goal is to improve the quality, informativity, and cost effectiveness of CNS post mortem examinations. A comprehensive study of the literature was conducted to provide a clinical background of neuropathological autopsy. There are numerous open questions in neuroscience, and new strategies are required to foster research in CNS diseases. These include the challenge of organizing brain banks tasked with managing and protecting detailed multidisciplinary information about their resources. Complex neuropathological analyses of post mortem series are also important to assess the effectiveness of diagnostics and therapy, identify environmental impact on the development of neurological disorders, and improve public health policy. The recommendations outline the need for collaboration between multiple specialists to establish the proper diagnosis and to broaden knowledge of neurological disorders.

MLK4 regulates DNA damage response and promotes triple-negative breast cancer chemoresistance.

Chemoresistance constitutes a major challenge in the treatment of triple-negative breast cancer (TNBC). Mixed-Lineage Kinase 4 (MLK4) is frequently amplified or overexpressed in TNBC where it facilitates the aggressive growth and migratory potential of breast cancer cells. However, the functional role of MLK4 in resistance to chemotherapy has not been investigated so far. Here, we demonstrate that MLK4 promotes TNBC chemoresistance by regulating the pro-survival response to DNA-damaging therapies. We observed that MLK4 knock-down or inhibition sensitized TNBC cell lines to chemotherapeutic agents in vitro. Similarly, MLK4-deficient cells displayed enhanced sensitivity towards doxorubicin treatment in vivo. MLK4 silencing induced persistent DNA damage accumulation and apoptosis in TNBC cells upon treatment with chemotherapeutics. Using phosphoproteomic profiling and reporter assays, we demonstrated that loss of MLK4 reduced phosphorylation of key DNA damage response factors, including ATM and CHK2, and compromised DNA repair via non-homologous end-joining pathway. Moreover, our mRNA-seq analysis revealed that MLK4 is required for DNA damage-induced expression of several NF-кB-associated cytokines, which facilitate TNBC cells survival. Lastly, we found that high MLK4 expression is associated with worse overall survival of TNBC patients receiving anthracycline-based neoadjuvant chemotherapy. Collectively, these results identify a novel function of MLK4 in the regulation of DNA damage response signaling and indicate that inhibition of this kinase could be an effective strategy to overcome TNBC chemoresistance.