New Insights into Therapy-Induced Progression of Cancer.

The malignant tumor is a complex heterogeneous set of cells functioning in a no less heterogeneous microenvironment. Like any dynamic system, cancerous tumors evolve and undergo changes in response to external influences, including therapy. Initially, most tumors are susceptible to treatment. However, remaining cancer cells may rapidly reestablish the tumor after a temporary remission. These new populations of malignant cells usually have increased resistance not only to the first-line agent, but also to the second- and third-line drugs, leading to a significant decrease in patient survival. Multiple studies describe the mechanism of acquired therapy resistance. In past decades, it became clear that, in addition to the simple selection of pre-existing resistant clones, therapy induces a highly complicated and tightly regulated molecular response that allows tumors to adapt to current and even subsequent therapeutic interventions. This review summarizes mechanisms of acquired resistance, such as secondary genetic alterations, impaired function of drug transporters, and autophagy. Moreover, we describe less obvious molecular aspects of therapy resistance in cancers, including epithelial-to-mesenchymal transition, cell cycle alterations, and the role of intercellular communication. Understanding these molecular mechanisms will be beneficial in finding novel therapeutic approaches for cancer therapy.

Exploring the diversity of cancer-associated fibroblasts: insights into mechanisms of drug resistance.

Introduction: Among the various stromal cell types within the tumor microenvironment, cancer-associated fibroblasts (CAFs) emerge as the predominant constituent, exhibiting a diverse array of oncogenic functions not intrinsic to normal fibroblasts. Their involvement spans across all stages of tumorigenesis, encompassing initiation, progression, and metastasis. Current understanding posits the coexistence of distinct subpopulations of CAFs within the tumor microenvironment across a spectrum of solid tumors, showcasing both pro- and antitumor activities. Recent advancements in single-cell transcriptomics have revolutionized our ability to meticulously dissect the heterogeneity inherent to CAF populations. Furthermore, accumulating evidence underscores the pivotal role of CAFs in conferring therapeutic resistance to tumors against various drug modalities. Consequently, efforts are underway to develop pharmacological agents specifically targeting CAFs. Methods: This review embarks on a comprehensive analysis, consolidating data from 36 independent single-cell RNA sequencing investigations spanning 17 distinct human malignant tumor types. Results: Our exploration centers on elucidating CAF population markers, discerning their prognostic relevance, delineating their functional contributions, and elucidating the underlying mechanisms orchestrating chemoresistance. Discussion: Finally, we deliberate on the therapeutic potential of harnessing CAFs as promising targets for intervention strategies in clinical oncology.

Therapy-induced secretion of spliceosomal components mediates pro-survival crosstalk between ovarian cancer cells.

Ovarian cancer often develops resistance to conventional therapies, hampering their effectiveness. Here, using ex vivo paired ovarian cancer ascites obtained before and after chemotherapy and in vitro therapy-induced secretomes, we show that molecules secreted by ovarian cancer cells upon therapy promote cisplatin resistance and enhance DNA damage repair in recipient cancer cells. Even a short-term incubation of chemonaive ovarian cancer cells with therapy-induced secretomes induces changes resembling those that are observed in chemoresistant patient-derived tumor cells after long-term therapy. Using integrative omics techniques, we find that both ex vivo and in vitro therapy-induced secretomes are enriched with spliceosomal components, which relocalize from the nucleus to the cytoplasm and subsequently into the extracellular vesicles upon treatment. We demonstrate that these molecules substantially contribute to the phenotypic effects of therapy-induced secretomes. Thus, SNU13 and SYNCRIP spliceosomal proteins promote therapy resistance, while the exogenous U12 and U6atac snRNAs stimulate tumor growth. These findings demonstrate the significance of spliceosomal network perturbation during therapy and further highlight that extracellular signaling might be a key factor contributing to the emergence of ovarian cancer therapy resistance.

Non-canonical functions of spliceosome components in cancer progression.

Dysregulation of pre-mRNA splicing is a common hallmark of cancer cells and it is associated with altered expression, localization, and mutations of the components of the splicing machinery. In the last few years, it has been elucidated that spliceosome components can also influence cellular processes in a splicing-independent manner. Here, we analyze open source data to understand the effect of the knockdown of splicing factors in human cells on the expression and splicing of genes relevant to cell proliferation, migration, cell cycle regulation, DNA repair, and cell death. We supplement this information with a comprehensive literature review of non-canonical functions of splicing factors linked to cancer progression. We also specifically discuss the involvement of splicing factors in intercellular communication and known autoregulatory mechanisms in restoring their levels in cells. Finally, we discuss strategies to target components of the spliceosome machinery that are promising for anticancer therapy. Altogether, this review greatly expands understanding of the role of spliceosome proteins in cancer progression.

Expression level of CD117 (KIT) on ovarian cancer extracellular vesicles correlates with tumor aggressiveness.

Ovarian cancer is known to be the most lethal malignancy among all gynecological cancers affecting a large number of women worldwide. The treatment of ovarian cancer is challenging due to the high recurrence rate of the disease and is further complicated by acquired chemoresistance. Most ovarian cancer deaths are the result of the metastatic spread of drug-resistant cells. The theory of cancer stem cells (CSC) suggests that both tumor initiation and progression are driven by a population of undifferentiated capable of self-renewal, tumor initiation and development of chemoresistance. The CD117 mast/stem cell growth factor receptor (KIT) is the most commonly used marker for ovarian CSCs. Here, we analyze the correlation between CD117 expression and histological tumor type in ovarian cancer cell lines (SK-OV-3 and MES-OV) and in small/medium extracellular vesicles (EVs) isolated from the urine of ovarian cancer patients. We have demonstrated that the abundance of CD117 on cells and EVs is correlated with tumor grade and therapy resistance status. Moreover, using small EVs isolated from ovarian cancer ascites, it was shown that recurrent disease is characterized by a much higher abundance of CD117 on EVs than primary tumor.

Deeper insights into transcriptional features of cancer-associated fibroblasts: An integrated meta-analysis of single-cell and bulk RNA-sequencing data.

Cancer-associated fibroblasts (CAFs) have long been known as one of the most important players in tumor initiation and progression. Even so, there is an incomplete understanding of the identification of CAFs among tumor microenvironment cells as the list of CAF marker genes varies greatly in the literature, therefore it is imperative to find a better way to identify reliable markers of CAFs. To this end, we summarized a large number of single-cell RNA-sequencing data of multiple tumor types and corresponding normal tissues. As a result, for 9 different types of cancer, we identified CAF-specific gene expression signatures and found 10 protein markers that showed strongly positive staining of tumor stroma according to the analysis of IHC images from the Human Protein Atlas database. Our results give an insight into selecting the most appropriate combination of cancer-associated fibroblast markers. Furthermore, comparison of different approaches for studying differences between cancer-associated and normal fibroblasts (NFs) illustrates the superiority of transcriptome analysis of fibroblasts obtained from fresh tissue samples. Using single-cell RNA sequencing data, we identified common differences in gene expression patterns between normal and cancer-associated fibroblasts, which do not depend on the type of tumor.

Alternative RNA splicing modulates ribosomal composition and determines the spatial phenotype of glioblastoma cells.

Glioblastoma (GBM) is characterized by exceptionally high intratumoral heterogeneity. However, the molecular mechanisms underlying the origin of different GBM cell populations remain unclear. Here, we found that the compositions of ribosomes of GBM cells in the tumour core and edge differ due to alternative RNA splicing. The acidic pH in the core switches before messenger RNA splicing of the ribosomal gene RPL22L1 towards the RPL22L1b isoform. This allows cells to survive acidosis, increases stemness and correlates with worse patient outcome. Mechanistically, RPL22L1b promotes RNA splicing by interacting with lncMALAT1 in the nucleus and inducing its degradation. Contrarily, in the tumour edge region, RPL22L1a interacts with ribosomes in the cytoplasm and upregulates the translation of multiple messenger RNAs including TP53. We found that the RPL22L1 isoform switch is regulated by SRSF4 and identified a compound that inhibits this process and decreases tumour growth. These findings demonstrate how distinct GBM cell populations arise during tumour growth. Targeting this mechanism may decrease GBM heterogeneity and facilitate therapy.

Chromatin Trapping of Factors Involved in DNA Replication and Repair Underlies Heat-Induced Radio- and Chemosensitization.

Hyperthermia has been used as an adjuvant treatment for radio- and chemotherapy for decades. In addition to its effects on perfusion and oxygenation of cancer tissues, hyperthermia can enhance the efficacy of DNA-damaging treatments such as radiotherapy and chemotherapy. Although it is believed that the adjuvant effects are based on hyperthermia-induced dysfunction of DNA repair systems, the mechanisms of these dysfunctions remain elusive. Here, we propose that elevated temperatures can induce chromatin trapping (c-trapping) of essential factors, particularly those involved in DNA repair, and thus enhance the sensitization of cancer cells to DNA-damaging therapeutics. Using mass spectrometry-based proteomics, we identified proteins that could potentially undergo c-trapping in response to hyperthermia. Functional analyses of several identified factors involved in DNA repair demonstrated that c-trapping could indeed be a mechanism of hyperthermia-induced transient deficiency of DNA repair systems. Based on our proteomics data, we showed for the first time that hyperthermia could inhibit maturation of Okazaki fragments and activate a corresponding poly(ADP-ribose) polymerase-dependent DNA damage response. Together, our data suggest that chromatin trapping of factors involved in DNA repair and replication contributes to heat-induced radio- and chemosensitization.

Therapy-induced stress response is associated with downregulation of pre-mRNA splicing in cancer cells.

BACKGROUND: Abnormal pre-mRNA splicing regulation is common in cancer, but the effects of chemotherapy on this process remain unclear. METHODS: To evaluate the effect of chemotherapy on slicing regulation, we performed meta-analyses of previously published transcriptomic, proteomic, phosphoproteomic, and secretome datasets. Our findings were verified by LC-MS/MS, western blotting, immunofluorescence, and FACS analyses of multiple cancer cell lines treated with cisplatin and pladienolide B. RESULTS: Our results revealed that different types of chemotherapy lead to similar changes in alternative splicing by inducing intron retention in multiple genes. To determine the mechanism underlying this effect, we analyzed gene expression in 101 cell lines affected by ɣ-irradiation, hypoxia, and 10 various chemotherapeutic drugs. Strikingly, оnly genes involved in the cell cycle and pre-mRNA splicing regulation were changed in a similar manner in all 335 tested samples regardless of stress stimuli. We revealed significant downregulation of gene expression levels in these two pathways, which could be explained by the observed decrease in splicing efficiency and global intron retention. We showed that the levels of active spliceosomal proteins might be further post-translationally decreased by phosphorylation and export into the extracellular space. To further explore these bioinformatics findings, we performed proteomic analysis of cisplatin-treated ovarian cancer cells. Finally, we demonstrated that the splicing inhibitor pladienolide B impairs the cellular response to DNA damage and significantly increases the sensitivity of cancer cells to chemotherapy. CONCLUSIONS: Decreased splicing efficiency and global intron retention is a novel stress response mechanism that may promote survival of malignant cells following therapy. We found that this mechanism can be inhibited by pladienolide B, which significantly increases the sensitivity of cancer cells to cisplatin which makes it a good candidate drug for improving the efficiency of cancer therapy.