Seminars in Oncology
Volume 39, Issue 1 , Pages 9-12, February 2012

Introduction: Molecular Pathogenesis of Hematologic Malignancies

  • Jaroslaw P. Maciejewski

      Affiliations

    • Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, Cleveland Clinic, Cleveland, OH
    • Corresponding Author InformationAddress correspondence to Jaroslaw P. Maciejewski, MD, PhD, FACP, Taussig Cancer Institute/R40 9500 Euclid Ave, Cleveland OH USA 44195
    • ,
  • Torsten Haferlach

      Affiliations

    • MLL Munich Leukemia Laboratory, Munich, Germany
    • Corresponding Author InformationProf Dr med, Dr phil Torsten Haferlach, MLL Munich Leukemia Laboratory, Max-Lebsche-Platz 31, 81377 Munich, Germany

Article Outline

 

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Future of Genomics in Hematologic Malignancies 

In addition to activating oncogenic mutations, deletions of chromosomal material, hypomorphic/inactivating mutations and aberrant promoter silencing of tumor-suppressor genes (TSG) have been recognized as important pathogenic mechanisms in myeloid malignancies, including acute myelogeneous leukemia (AML), myelodysplastic syndrome (MDS), and myeloproliferative neoplasms (MPN), as well as overlap entities between these diseases (MPN/MDS). Recently, tremendous progress has been made in the molecular investigations of the pathogenesis of these diseases, largely due to broad application of high-throughput molecular technologies, including single-nucleotide polymorphism arrays (SNP-A), comparative genomic hybridization arrays (CGH-A), and especially whole-genome next-generation sequencing (NGS).1, 2, 3

One could envision that in the nearest future most of the pathogenetic somatic mutations occurring at the frequencies of greater than 1% of all major types of leukemia and other hematologic malignancies will be identified and subsequent work will focus on the mechanistic studies. These investigations will have to include the contribution of the identified lesions to the leukemogenesis, clarification of tremendous heterogeneity and genotype/phenotype relationships, distinction between ancestral and secondary lesions, identification of drugable targets, and the assignment of clinical (diagnostic/prognostic) relevance of these defects. The latter task will require selection of clinically applicable techniques for routine molecular diagnostics and incorporation of molecular information in the diagnostic algorithms and prognostic schemes. Future studies also will have to address characterization of methylome and methyhydroxylome, both pathologic as well as normal, in a tissue-specific manner.

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Clinical Diversity as a Reflection of Molecular Heterogeneity 

Whole-genome NGS and systematic targeted sequencing projects1 led to realization that in many malignancies patterns of mutations can be found; this underlying molecular diversity likely translates into clinical heterogeneity.4, 5, 6 Similarly, various seemingly unrelated molecular events may produce a biologic phenocopy through molecular pathway divergence and through substitution of defects. For example, activating mutations in the PTK/JAK/STAT pathway can be substituted by defects in signal deactivation through defective phosphatases or ubiquitin ligases.7 Similarly, both hypomorphic/inactivating heterozygous mutations in TSGs can lead to haplo-insufficiency,8 but analogous decrease in the availability of intact mRNA may be due to spliceosomal defects resulting from missplicing and subsequent missense-mediated decay9 (see also below).

The heterogeneity of clinical presentation, morphology and outcomes also may be related to the types and topography of the molecular defects. In many instances and in contrast to activating mutations, canonical mutations are not present and instead the locations of mutations and their type (such as mis/nonsense and frame-shift mutations) may correspond to variable severity and speed of progression. Similarly, some mutations occur in heterozygous while others in hemizygous or even homozygous configurations through somatic uniparental disomy (sUPD). Somatic deletion and sUPD can affect a large number of allelic polymorphisms, rendering the mutant cells hemizygous or homozygous for germline-encoded hypomorphic variants and thereby affecting the resultant phenotype.8 Similarly, there are potential interactions between methylation and hydroxymethylation events and sUPD, whereby, in the areas of hemimethylation, the retained and duplicated allele may be methylated and thus repressed, or conversely the silenced allele may be deleted (Figure 1).

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Groups of Genes Affected by Mutations 

Conceptually, somatic mutations target genes with similar and complementary function affect members of common pathways. To facilitate mechanistic insights into the pathologic process, they can be grouped based on their functional characteristics and pathological consequences (Figure 1). Of course there are many other ways to categorize somatic mutations, including those that are ancestral (type I) versus facilitator mutations (type II), those that affect the stem cell function (maintenance, survival) versus those that alter progenitor cells (proliferation, differentiation), or according to their pathomorphologic or genetic context (eg, secondary AML [sAML] v MDS/MPN, juvenile myelomonocytic leukemia [JMML] v chronic myelomonocytic leukemia [CMML]) in conjunction with complex karyotype v single numerical alterations. Examples of groups of genes categorized by functional characteristics or by pathway effects are included below.

Activating Proliferative Mutations 

Various activating mutations have been identified in hematologic malignancies and include phosphothyrosine kinase receptors (TRKs) such as cKIT, cMPL, and FLT3, but also transduction proteins subject to phosphorylation such as JAK2 or proteins involved in deactivation of activation signals transduced by phosphorylated protein kinases such as CBL, an E3 ubiquitine ligase that can inactivate a number of phophorylated targets. These mutations can occur in heterozygous forms and sUPD may lead to homozygozity as a sign of progression, unlike mutations in some TSGs, which are not associated with hemizygous deletions.7

Gene Mutations Affecting Differentiation 

Some TSGs can affect differentiation and thereby more or less directly survival. Examples of such signal transduces that are frequently effected by mutations are CEBPA, RUNX1, MLL, GATA1, and GATA2 and likely NPM1. These genes can be mutated in various configurations, including homozygous forms, but are often haplo-insufficient in cases with the wild-type of these genes.

Antiapoptotic Tumor Suppressor Gene Mutations 

Classical TSGs are important contributors to leukemic progression. Classical examples of such gene mutations with clear clinical impact include TP53, MDM2, and MDM4.

Genes Involved in Epigenetic Regulation 

In myeloid malignancies, genomic instability has been used as a concept explaining genomic lesions (including chromosomal abnormalities and in a broader context also somatic mutations) beyond their random appearance. A great deal of work has been devoted to the study of various pathways of DNA repair machinery. In addition to the mostly irreversible genomic lesions, epigenetic changes are believed to be a hallmark of malignant evolution. DNA repair pathways may be linked to aberrant methylation status of CpG islands (including aberrant hyper- and hypomethylations), and chromosomal aberrations observed in cancer may be related to processes of DNA repair. However, defective methylation, including hypo- and hypermethylation of promoter DNA, relates to the concept of epigenetic instability. Discovery of somatic mutations in various epigenetic regulatory genes may constitute a link between the genomic and epigenetic instability. Mutations in IDH1/2, resulting in production of the neomorphic oncogenic substrate oxoglutyrate, are linked functionally to TET2 mutations and result in decreased methyl-hydroxymethylation.10, 11, 12 Mutations in DNMT3A seem to indirectly affect related pathways but are not mutually exclusive and the pathogeneic link to TET2 and IDH2 mutations has not been established. Another group of “epigenetic” mutations includes EZH2, SUS12, EED, ASXL1, and UTX and affects methylation of various histone marks.13, 14, 15

Mutations of Spliceosomal Genes 

Unbiased, whole-genome sequencing approaches led to identification of a large number of new somatic mutations in myeloid malignancies. Recently, we and others have discovered somatic mutations in a gene coding spliceosomal protein, SF3B1, associated with a distinct form of MDS characterized by the presence of ring sideroblasts.16 This finding focused the search for molecular lesions involving U2 spliceosome complex and somatic mutations have been identified in other important members of spliceosome machinery (to date, mutations in 15 spliceosomal genes have been described), including U2AF1, SRSF2, and ZRSR2.9, 16, 17 They are associated with different clinical phenotypes and pathomorphology; SF3B1 are present in low-grade MDS with ring sideroblasts, while U2AF1, SRSF2, and ZRSR2 are particularly frequent in CMML and advanced forms of MDS, but also in AML. These mutations point to a new important and ubiquitous leukemogenic pathway and thereby spliceosomal genes are likely bona fide tumor suppressors. Mutations in these genes may constitute diagnostic biomarkers and will serve as targets of future therapies. The occurrence of mutations of spliceosomal proteins involved in various stages of this process suggests that the splicing process is vulnerable to acquisition of molecular defects. In our studies in myeloid malignancies, the most commonly affected spliceosomal genes were SF3B1, U2AF1, and SFSR2, but likely other hematologic and solid tumors can harbor mutations of diverse spliceosomal genes.

Mechanistically defective splicing may have similar consequences as loss of function mutations through retention of introns. Similarly, nonsense-induced decay may lead to removal of defective transcripts and result in overall decreased expression of the corresponding genes. Deep sequencing of RNA in hematopoietic cells derived from mutant cases suggests that mis-splicing or exon skipping rather than abnormal alternative splicing is the consequence of mutations.9 The splicing defects resulting from mutations likely affect a selected subset of TSGs and lead to their functional haplo-insuffciency with clinical features similar to other lesions affecting corresponding target genes (Figure 2).

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Mutational Burden, Clonal Size 

Novel technologies including ability to perform deep sequencing and easy distinction of hemizygous versus heterozygous or homozygous lesions may add another layer of complexity in the interpretation of molecular data. They allow not only for distinction of these types of lesions from varying impact of contamination with nonclonal cells but also for determination of mutational burden. The impact of clonal size/mutational load has been explored in target fashion for many lesions such as JAK2 mutations, but deep sequencing of many genes brings about realization of subclonal evolution and succession of clonal dominance in the course of disease (Figure 3). Clonal dynamics and mutational burden may further help to substratify the prognostic algorithms.18 Similarly, it is likely that presence of tiny subclones harboring prognostically unfavorable defects may be of clinical impact.

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Conclusions 

Mechanistically, many types of lesion converge into common downstream effects on a subsets of target genes that are involved in fundamental cellular processes. The relationship may be functional as seen, for example, in similar proliferative effects of CBL and TKRs or inhibition of methylcytosine hydroxylation IDH1/2 and TET2 or histone methylation IDH1/2 and UTX. On the genetic level, diverse types of defects affecting specific genes may generate a phencopy; for example, heterozygous hypomorphic/inactivating TSG mutations, spliceosomal mutations affecting splicing of TSGs, or epigenetic silencing lead to haplo-insufficient gene expression (Figure 2). Ongoing progress in molecular technologies will be applied to the diagnostic routine as it is the case for CGH, SNP-A–based karyotyping and capture, and NGS-based technologies. They all are likely to be adopted to the clinical need and lead to introduction of multi-analytic diagnostic tests allowing for testing of not only canonical mutations but screening of whole genes recurrently affected by mutations as well. This approach will likely revolutionize molecular diagnostic of cancer and lead to personalized therapeutic approaches. Molecular tests including the presence of certain somatic mutations or their combinations will likely affect newer prognostic schemes and possible combination of mutations may define nosologic subentities rather than morphologic criteria, which often do not correspond to common pathogenetic pathways. The selection of articles in this issue of Seminars in Oncology illustrates some of the most important aspects of cancer genetics as it applies to hematologic malignancies.

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References 

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PII: S0093-7754(11)00303-4

doi:10.1053/j.seminoncol.2011.12.002

Seminars in Oncology
Volume 39, Issue 1 , Pages 9-12, February 2012

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