Engineered ECM models: Opportunities to advance understanding of tumor heterogeneity

Intratumoral heterogeneity is a negative prognostic factor for cancer and commonly attributed to microenvironment-driven genetic mutations and/or the emergence of cancer stem-like cells. How aberrant extracellular matrix (ECM) remodeling regulates the phenotypic diversity of tumor cells, however, remains poorly understood due in part to a lack of model systems that allow isolating the physicochemical heterogeneity of malignancy-associated ECM for mechanistic studies. Here, we review the compositional, microarchitectural, and mechanical hallmarks of cancer-associated ECM and highlight biomaterials and engineering approaches to recapitulate these properties for in vitro and in vivo studies. Subsequently, we describe how such engineered platforms may be explored to define the spatiotemporal dynamics through which cancer-associated ECM remodeling regulates intratumoral heterogeneity and the cancer stem-like cell phenotype. Finally, we highlight future opportunities and technological advances to further elucidate the relationship between tumor-associated ECM dynamics and intratumoral heterogeneity.     

Modeling somatic evolution in tumorigenesis

Tumorigenesis in humans is thought to be a multistep process where certain mutations confer a selective advantage, allowing lineages derived from the mutated cell to outcompete other cells. Although molecular cell biology has substantially advanced cancer research, our understanding of the evolutionary dynamics that govern tumorigenesis is limited. This paper analyzes the computational implications of cancer progression presented by Hanahan and Weinberg in The Hallmarks of Cancer. We model the complexities of tumor progression as a small set of underlying rules that govern the transformation of normal cells to tumor cells. The rules are implemented in a stochastic multistep model. The model predicts that (i) early-onset cancers proceed through a different sequence of mutation acquisition than late-onset cancers; (ii) tumor heterogeneity varies with acquisition of genetic instability, mutation pathway, and selective pressures during tumorigenesis; (iii) there exists an optimal initial telomere length which lowers cancer incidence and raises time of cancer onset; and (iv) the ability to initiate angiogenesis is an important stage-setting mutation, which is often exploited by other cells. The model offers insight into how the sequence of acquired mutations affects the timing and cellular makeup of the resulting tumor and how the cellular-level population dynamics drive neoplastic evolution.     

Hormone resistance and neuroendocrine differentiation due to accumulation of genetic lesions during clonal evolution of prostate cancer

Progression of malignant tumors is largely due to clonal evolution of the primary tumor, clones acquiring different sets of molecular genetic lesions. Lesions can confer a selective advantage in proliferation rate or metastasis on the tumor cell population, especially if developing resistance to anticancer therapy. Prostate cancer (PCa) provides an illustrative example of clinically significant clonal evolution. The review considers the genetic alterations that occur in primary PCa and the mechanism whereby hormone-refractory PCa develops on hormone therapy, including mutations and alternative splicing of the androgen receptor gene (AR) and intratumoral androgen synthesis. Certain molecular genetic lesions determine resistance to new generation inhibitors (AR mutations that block the antagonist effect or allow other hormones to activate the receptor) or lead to neuroendocrine differentiation (repression of the AR signaling pathway, TP53 mutations, and amplification of the AURKA or MYCN oncogene). Multistep therapy based on the data about somatic mutations associated with progression and metastasis of the primary tumor can be expected to significantly improve the survival of patients with advanced PCa in the nearest future.     

The Malignant Pleural Effusion as a Model to Investigate Intratumoral Heterogeneity in Lung Cancer

Malignant Pleural Effusions (MPE) may be useful as a model to study hierarchical progression of cancer and/or intratumoral heterogeneity. To strengthen the rationale for developing the MPE-model for these purposes, we set out to find evidence for the presence of cancer stem cells (CSC) in MPE and demonstrate an ability to sustain intratumoral heterogeneity in MPE-primary cultures. Our studies show that candidate lung CSC-expression signatures (PTEN, OCT4, hTERT, Bmi1, EZH2 and SUZ12) are evident in cell pellets isolated from MPE, and MPE-cytopathology also labels candidate-CSC (CD44, cMET, MDR-1, ALDH) subpopulations. Moreover, in primary cultures that use MPE as the source of both tumor cells and the tumor microenvironment (TME), candidate CSC are maintained over time. This allows us to live-sort candidate CSC-fractions from the MPE-tumor mix on the basis of surface markers (CD44, c-MET, uPAR, MDR-1) or differences in xenobiotic metabolism (ALDH). Thus, MPE-primary cultures provide an avenue to extract candidate CSC populations from individual (isogenic) MPE-tumors. This will allow us to test whether these cells can be discriminated in functional bioassays. Tumor heterogeneity in MPE-primary cultures is evidenced by variable immunolabeling, differences in colony-morphology, and differences in proliferation rates of cell subpopulations. Collectively, these data justify the ongoing development of the MPE-model for the investigation of intratumoral heterogeneity, tumor-TME interactions, and phenotypic validation of candidate lung CSC, in addition to providing direction for the pre-clinical development of rational therapeutics.     

Niche-Dependent Gene Expression Profile of Intratumoral Heterogeneous Ovarian Cancer Stem Cell Populations

Intratumoral heterogeneity challenges existing paradigms for anti-cancer therapy. We have previously demonstrated that the human embryonic stem cells (hESC)-derived cellular microenvironment in immunocompromised mice, enables functional distinction of heterogeneous tumor cells, including cells which do not grow into a tumor in a conventional direct tumor xenograft platform. We have identified and characterized six cancer cell subpopulations each clonally expanded from a single cell, derived from human ovarian clear cell carcinoma of a single tumor, to demonstrate striking intratumoral phenotypic heterogeneity that is dynamically dependent on the tumor growth microenvironment. These cancer cell subpopulations, characterized as cancer stem cell subpopulations, faithfully recapitulate the full spectrum of histological phenotypic heterogeneity known for human ovarian clear cell carcinoma. Each of the six subpopulations displays a different level of morphologic and tumorigenic differentiation wherein growth in the hESC-derived microenvironment favors growth of CD44+/aldehyde dehydrogenase positive pockets of self-renewing cells that sustain tumor growth through a process of tumorigenic differentiation into CD44-/aldehyde dehydrogenase negative derivatives. Strikingly, these derivative cells display microenvironment-dependent plasticity with the capacity to restore self-renewal markers and CD44 expression. In the current study, we delineate the distinct gene expression and epigenetic profiles of two such subpopulations, representing extremes of phenotypic heterogeneity in terms of niche-dependent self-renewal and tumorigenic differentiation. By combining Gene Set Enrichment, Gene Ontology and Pathway-focused array analyses with methylation status, we propose a suite of robust differences in tumor self-renewal and differentiation pathways that underlie the striking intratumoral phenotypic heterogeneity which characterize this and other solid tumor malignancies.     

Establishment and Characterization of a Highly Tumourigenic and Cancer Stem Cell Enriched Pancreatic Cancer Cell Line as a Well Defined Model System

Standard cancer cell lines do not model the intratumoural heterogeneity situation sufficiently. Clonal selection leads to a homogeneous population of cells by genetic drift. Heterogeneity of tumour cells, however, is particularly critical for therapeutically relevant studies, since it is a prerequisite for acquiring drug resistance and reoccurrence of tumours. Here, we report the isolation of a highly tumourigenic primary pancreatic cancer cell line, called JoPaca-1 and its detailed characterization at multiple levels. Implantation of as few as 100 JoPaca-1 cells into immunodeficient mice gave rise to tumours that were histologically very similar to the primary tumour. The high heterogeneity of JoPaca-1 was reflected by diverse cell morphology and a substantial number of chromosomal aberrations. Comparative whole-genome sequencing of JoPaca-1 and BxPC-3 revealed mutations in genes frequently altered in pancreatic cancer. Exceptionally high expression of cancer stem cell markers and a high clonogenic potential in vitro and in vivo was observed. All of these attributes make this cell line an extremely valuable model to study the biology of and pharmaceutical effects on pancreatic cancer.     

Heterogeneity of KRAS Mutation Status in Rectal Cancer

IntroductionAnti-EGFR targeted therapy is of increasing importance in advanced colorectal cancer and prior KRAS mutation testing is mandatory for therapy. However, at which occasions this should be performed is still under debate. We aimed to assess in patients with locally advanced rectal cancer whether there is intra-specimen KRAS heterogeneity prior to and upon preoperative chemoradiotherapy (CRT), and if there are any changes in KRAS mutation status due to this intervention.ResultsFor 20 (43%) out of the 47 patients, a KRAS mutation was detected. With 12 out of 20, the majority of these mutations affected codon 35. We did not obtained evidence that CRT results in changes of the KRAS mutation pattern. In addition, no intratumoral heterogeneity in the KRAS mutational status could be proven. This was true for both the biopsies prior to CRT and the resection specimens thereafter. The discrepancy observed in some samples when using the SNaPshot™ assay was due to insufficient sensitivity of this technique upon massive tumor regression by CRT as application of the therascreen® KRAS test revealed concordant results.ConclusionOur results indicate that the KRAS mutation status at the primary tumor site of rectal cancer is homogenous. Its assessment for therapeutic decisions is feasible in pre-therapeutic biopsies as well as in post-therapeutic resected specimens. The amount of viable tumor cells seems to be an important determinant for assay sensitivity and should thus be considered for selection of the analytical method.     

Clonal Analysis in Recurrent Astrocytic, Oligoastrocytic and Oligodendroglial Tumors Implicates IDH1- Mutation as Common Tumor Initiating Event

Background

To investigate the dynamics of inter- and intratumoral molecular alterations during tumor progression in recurrent gliomas.

Methodology/Principal Findings

To address intertumoral heterogeneity we investigated non- microdissected tumor tissue of 106 gliomas representing 51 recurrent tumors. To address intratumoral heterogeneity a set of 16 gliomas representing 7 tumor pairs with at least one recurrence, and 4 single mixed gliomas were investigated by microdissection of distinct oligodendroglial and astrocytic tumor components. All tumors and tumor components were analyzed for allelic loss of 1p/19q (LOH 1p/19q), for TP53- mutations and for R132 mutations in the IDH1 gene. The investigation of non- microdissected tumor tissue revealed clonality in 75% (38/51). Aberrant molecular alterations upon recurrence were detected in 25% (13/51). 64% (9/14) of these were novel and associated with tumor progression. Loss of previously detected alterations was observed in 36% (5/14). One tumor pair (1/14; 7%) was significant for both. Intratumoral clonality was detected in 57% (4/7) of the microdissected tumor pairs and in 75% (3/4) of single microdissected tumors. 43% (3/7) of tumor pairs and one single tumor (25%) revealed intratumoral heterogeneity. While intratumoral heterogeneity affected both the TP53- mutational status and the LOH1p/19q status, all tumors with intratumoral heterogeneity shared the R132 IDH1- mutation as a common feature in both their microdissected components.

Conclusions/Significance

The majority of recurrent gliomas are of monoclonal origin. However, the detection of divertive tumor cell clones in morphological distinct tumor components sharing IDH1- mutations as early event may provide insight into the tumorigenesis of true mixed gliomas.     

Acute myeloid leukaemia: a paradigm for the clonal evolution of cancer?

Acute myeloid leukaemia (AML) is an uncontrolled clonal proliferation of abnormal myeloid progenitor cells in the bone marrow and blood. Advances in cancer genomics have revealed the spectrum of somatic mutations that give rise to human AML and drawn our attention to its molecular evolution and clonal architecture. It is now evident that most AML genomes harbour small numbers of mutations, which are acquired in a stepwise manner. This characteristic, combined with our ability to identify mutations in individual leukaemic cells and our detailed understanding of normal human and murine haematopoiesis, makes AML an excellent model for understanding the principles of cancer evolution. Furthermore, a better understanding of how AML evolves can help us devise strategies to improve the therapy and prognosis of AML patients. Here, we draw from recent advances in genomics, clinical studies and experimental models to describe the current knowledge of the clonal evolution of AML and its implications for the biology and treatment of leukaemias and other cancers.KEY WORDS: Acute myeloid leukaemia, Cancer, Clonal evolution, In vivo models of leukaemia, Mutation     

Natural and chemotherapy-induced clonal evolution of tumors

Evolution and natural selection of tumoral clones in the process of transformation and the following carcinogenesis can be called natural clonal evolution. Its main driving factors are internal: genetic instability initiated by driver mutations and microenvironment, which enables selective pressure while forming the environment for cell transformation and their survival. We present our overview of contemporary research dealing with mechanisms of carcinogenesis in different localizations from precancerous pathologies to metastasis and relapse. It shows that natural clonal evolution establishes intratumoral heterogeneity and enables tumor progression. Tumors of monoclonal origin are of low-level intratumoral heterogeneity in the initial stages, and this increases with the size of the tumor. Tumors of polyclonal origin are of extremely high-level intratumoral heterogeneity in the initial stages and become more homogeneous when larger due to clonal expansion. In cases of chemotherapy-induced clonal evolution of a tumor, chemotherapy becomes the leading factor in treatment. The latest research shows that the impact of chemotherapy can radically increase the speed of clonal evolution and lead to new malignant and resistant clones that cause tumor metastasis. Another option of chemotherapy-induced clonal evolution is formation of a new dominant clone from a clone that was minor in the initial tumor and obtained free space due to elimination of sensitive clones by chemotherapy. As a result, in ~20% of cases, chemotherapy can stimulate metastasis and relapse of tumors due to clonal evolution. The conclusion of the overview formulates approaches to tumor treatment based on clonal evolution: in particular, precision therapy, prediction of metastasis stimulation in patients treated with chemotherapy, methods of genetic evaluation of chemotherapy efficiency and clonal-oriented treatment, and approaches to manipulating the clonal evolution of tumors are presented.     

Characterization of a novel microfluidic platform for the isolation of rare single cells to enable CTC analysis from head and neck squamous cell carcinoma patients

Detailed examination of tumor components is leading‐edge to establish personalized cancer therapy. Accompanying research on cell‐free DNA, the cell count of circulating tumor cells (CTCs) in patient blood is seen as a crucial prognostic factor. The potential of CTC analysis is further not limited to the determination of the overall survival rate but sheds light on understanding inter‐ and intratumoral heterogeneity. In this regard, commercial CTC isolation devices combining an efficient enrichment of rare cells with a droplet deposition of single cells for downstream analysis are highly appreciated. The Liquid biopsy platform CTCelect was developed to realize a fully‐automated enrichment and single cell dispensing of CTCs from whole blood without pre‐processing. We characterized each process step with two different carcinoma cell lines demonstrating up to 87 % enrichment (n = 10) with EpCAM coupled immunomagnetic beads, 73 % optical detection and dispensing efficiency (n = 5). 40 to 56.7 % of cells were recovered after complete isolation from 7.5 ml untreated whole blood (n = 6). In this study, CTCelect enabled automated dispensing of single circulating tumor cells from HNSCC patient samples, qPCR‐based confirmation of tumor‐related biomarkers and immunostaining. Finally, the platform was compared to commercial CTC isolation technologies to highlight advantages and limitations of CTCelect. This system offers new possibilities for single cell screening in cancer diagnostics, individual therapy approaches and real‐time monitoring.     

肿瘤遗传异质性

尽管大多数自发性肿瘤源于单细胞, 但人们逐渐认识到个体肿瘤的内部存在异质性, 这种差异涉及分化程度、细胞增殖率、侵袭和转移能力及治疗反应等众多方面。分子生物学研究也证实肿瘤演进过程中不断产生新的突变, 为肿瘤内部存在异质性增加了更有力的佐证。文章综述了关于肿瘤遗传异质性研究的主要进展。鉴于遗传异质性分析可为揭示肿瘤细胞的产生、扩散、转移的时间段提供重要信息, 文章以瘤内异质性为主线, 列举了遗传异质性存在的实验依据; 阐述了遗传多样性在人类个体肿瘤发展进化史中的研究价值; 介绍了遗传异质性产生的两种模式, 包括肿瘤干细胞模型和克隆进化模型; 总结了遗传异质性在肿瘤转移、治疗中的重要临床意义。文章最后展示了常用的遗传异质性的研究方法, 包括特定基因的分析方法和基于基因组水平的方法, 并对各自的优缺点进行了分析。     

Comparative Oncogenomics Implicates the Neurofibromin 1 Gene (NF1) as a Breast Cancer Driver

Identifying genomic alterations driving breast cancer is complicated by tumor diversity and genetic heterogeneity. Relevant mouse models are powerful for untangling this problem because such heterogeneity can be controlled. Inbred Chaos3 mice exhibit high levels of genomic instability leading to mammary tumors that have tumor gene expression profiles closely resembling mature human mammary luminal cell signatures. We genomically characterized mammary adenocarcinomas from these mice to identify cancer-causing genomic events that overlap common alterations in human breast cancer. Chaos3 tumors underwent recurrent copy number alterations (CNAs), particularly deletion of the RAS inhibitor Neurofibromin 1 (Nf1) in nearly all cases. These overlap with human CNAs including NF1, which is deleted or mutated in 27.7% of all breast carcinomas. Chaos3 mammary tumor cells exhibit RAS hyperactivation and increased sensitivity to RAS pathway inhibitors. These results indicate that spontaneous NF1 loss can drive breast cancer. This should be informative for treatment of the significant fraction of patients whose tumors bear NF1 mutations.     

Detection and Clinical Significance of Intratumoral EGFR Mutational Heterogeneity in Chinese Patients with Advanced Non-Small Cell Lung Cancer

Purpose

This study evaluated occurrence and potential clinical significance of intratumoral EGFR mutational heterogeneity in Chinese patients with non-small cell lung cancer (NSCLC).

Materials and Methods

Eighty-five stage IIIa-IV NSCLC patients who had undergone palliative surgical resection were included in this study. Of these, 45 patients carried EGFR mutations (group-M) and 40 patients were wild-type (group-W). Each tumor sample was microdissected to yield 28–34 tumor foci and Intratumoral EGFR mutation were determined using Denaturing High Performance Liquid Chromatography (DHPLC) and Amplification Refractory Mutation System (ARMS). EGFR copy numbers were measured using fluorescence in situ hybridization (FISH).

Results

Microdissection yielded 1,431 tumor foci from EGFR mutant patients (group-M) and 1,238 foci from wild-type patients (group-W). The EGFR mutant frequencies in group-M were 80.6% (1,154/1,431) and 87.1% (1,247/1,431) using DHPLC and ARMS, respectively. A combination of EGFR-mutated and wild-type cells was detected in 32.9% (28/85) of samples by DHPLC and 28.2% (24/85) by ARMS, supporting the occurrence of intratumoral heterogeneity. Thirty-one patients (36.5%) were identified as EGFR FISH-positive. Patients harboring intratumoral mutational heterogeneity possessed lower EGFR copy numbers than those tumors contained mutant cells alone (16.7% vs. 71.0%, P<0.05). Among 26 patients who had received EGFR-TKIs, the mean EGFR mutation content was higher in patients showing partial response (86.1%) or stable disease (48.7%) compared with patients experiencing progressive disease (6.0%) (P = 0.001). There also showed relationship between progression-free survival (PFS) and different content of EGFR mutation groups (pure wild type EGFR, EGFR mutation with heterogeneity and pure mutated EGFR) (P = 0.001).

Conclusion

Approximately 30% of patients presented intratumoral EGFR mutational heterogeneity, accompanying with relatively low EGFR copy number. EGFR mutant content was correlated with the response and prognosis of EGFR-TKIs.     

Evolution and Phenotypic Selection of Cancer Stem Cells

Cells of different organs at different ages have an intrinsic set of kinetics that dictates their behavior. Transformation into cancer cells will inherit these kinetics that determine initial cell and tumor population progression dynamics. Subject to genetic mutation and epigenetic alterations, cancer cell kinetics can change, and favorable alterations that increase cellular fitness will manifest themselves and accelerate tumor progression. We set out to investigate the emerging intratumoral heterogeneity and to determine the evolutionary trajectories of the combination of cell-intrinsic kinetics that yield aggressive tumor growth. We develop a cellular automaton model that tracks the temporal evolution of the malignant subpopulation of so-called cancer stem cells(CSC), as these cells are exclusively able to initiate and sustain tumors. We explore orthogonal cell traits, including cell migration to facilitate invasion, spontaneous cell death due to genetic drift after accumulation of irreversible deleterious mutations, symmetric cancer stem cell division that increases the cancer stem cell pool, and telomere length and erosion as a mitotic counter for inherited non-stem cancer cell proliferation potential. Our study suggests that cell proliferation potential is the strongest modulator of tumor growth. Early increase in proliferation potential yields larger populations of non-stem cancer cells(CC) that compete with CSC and thus inhibit CSC division while a reduction in proliferation potential loosens such inhibition and facilitates frequent CSC division. The sub-population of cancer stem cells in itself becomes highly heterogeneous dictating population level dynamics that vary from long-term dormancy to aggressive progression. Our study suggests that the clonal diversity that is captured in single tumor biopsy samples represents only a small proportion of the total number of phenotypes.     

Cancer stem cell plasticity and tumor hierarchy

The origins of the complex process of intratumoral heterogeneity have been highly debated and different cellular mechanisms have been hypothesized to account for the diversity within a tumor. The clonal evolution and cancer stem cell(CSC) models have been proposed as drivers of this heterogeneity. However, the concept of cancer stem cell plasticity and bidirectional conversion between stem and non-stem cells has added additional complexity to these highly studied paradigms and may help explain the tumor heterogeneity observed in solid tumors. The process of cancer stem cell plasticity in which cancer cel s harbor the dynamic ability of shifting from a non-CSC state to a CSC state and vice versa may be modulated by specific microenvironmental signals and cellular interactions arising in the tumor niche. In addition to promoting CSC plasticity, these interactions may contribute to the cellular transformation of tumor cells and affect response to chemotherapeutic and radiation treatments by providing CSCs protection from these agents. Herein, we review the literature in support of this dynamic CSC state, discuss the effectors of plasticity, and examine their role in the development and treatment of cancer.     

Intratumor heterogeneity and clonal evolution revealed in castration-resistant prostate cancer by longitudinal genomic analysis

Intratumor heterogeneity is a key driver for local relapse and treatment failure. Thus, using multifocal prostate cancer as a model to investigate tumor inter-clonal relationships and tumor evolution could aid in our understanding of drug resistance. Previous studies discovered genomic alterations by comparing hormone-sensitive prostate cancer (HSPC) with castration-resistant prostate cancer (CRPC) in large cohorts. However, most studies did not sequentially sample tumors from the same patient. In our study, we performed whole-exome sequencing (WES) on 14 specimens from five locally relapsed patients before and after androgen-deprivation therapy. We described the landscape of genomic alterations before and after treatment and identified critical driver events that could have contributed to the evolution of CRPC. In addition to confirming known cancer genes such as TP53 and CDK12, we also identified new candidate genes that may play a role in the progression of prostate cancer, including MYO15A, CHD6 and LZTR1. At copy number alteration (CNA) level, gain of 8q24.13-8q24.3 was observed in 60% of patients and was the most commonly altered locus in both HSPC and CRPC tumors. Finally, utilizing phylogenetic reconstruction, we explored the clonal progression pattern from HSPC to CRPC in each patient. Our findings highlight the complex and heterogeneous mechanisms underlying the development of drug resistance, and underscore the potential value of monitoring tumor clonal architectures during disease progression in a clinical setting.     

A general framework for analyzing tumor subclonality using SNP array and DNA sequencing data

Intra-tumor heterogeneity reflects cancer genome evolution and provides key information for diagnosis and treatment. When bulk tumor tissues are profiled for somatic copy number alterations (sCNA) and point mutations, it may be difficult to estimate their cellular fractions when a mutation falls within a sCNA. We present the Clonal Heterogeneity Analysis Tool, which estimates cellular fractions for both sCNAs and mutations, and uses their distributions to inform macroscopic clonal architecture. In a set of approximately 700 breast tumors, more than half appear to contain multiple recognizable aneuploid tumor clones, and many show subtype-specific differences in clonality for known cancer genes.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-014-0473-4) contains supplementary material, which is available to authorized users.     

Structural Alterations in Human Fibroblast Growth Factor Receptors in Carcinogenesis

Fibroblast growth factor (FGF) plays an important role in human embryogenesis, angiogenesis, cell proliferation, and differentiation. Carcinogenesis is accompanied by aberrant constitutive activation of FGF receptors (FGFRs) resulting from missense mutation in the FGFR1-4 genes, generation of chimeric oncogenes, FGFR1-4 gene amplification, alternative splicing shift toward formation of mesenchymal FGFR isoforms, and FGFR overexpression. Altogether, these alterations contribute to auto-and paracrine stimulation of cancer cells and neoangiogenesis. Certain missense mutations are found at a high rate in urinary bladder cancer and can be used for non-invasive cancer recurrence diagnostics by analyzing urine cell pellet DNA. Chimeric FGFR1/3 and amplified FGFR1/2 genes can predict cell response to the targeted therapy in various oncological diseases. In recent years, high-throughput sequencing has been used to analyze exomes of virtually all human tumors, which allowed to construct phylogenetic trees of clonal cancer evolution with special emphasis on driver mutations in FGFR1-4 genes. At present, FGFR blockers, such as multi-kinase inhibitors, specific FGFR inhibitors, and FGF ligand traps are being tested in clinical trials. In this review, we discuss current data on the functioning of the FGFR family proteins in both normal and cancer cells, mutations in the FGFR1-4 genes, and mechanisms underlying their oncogenic potential, which might be interesting to a broad range of scientists searching for specific tumor markers and targeted anti-cancer drugs.