No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
Chronic myelocytic leukemia: Clonal origin in a stem cell common to the granulocyte, erythrocyte, platelet and monocyte/macrophage
Abstract
Glucose-6-phosphate dehydrogenase (G-6-PD) isoenzymes types of granulocytes were determined in eight women with chronic myelocytic leukemia (CML). The patients were heterozygous at the X-linked G-6-PD locus for the common gene, GdB, and a variant, such as GdA, so that both B and A enzyme types were found in skin cells. In contrast to these normal cells, only one G-6-PD type was found in CML granulocytes. The fact that such single-enzyme phenotypes are found in CML granulocytes, but not in nonleukemic granulocytes, provides strong evidence that the disease has a clonal origin. Single-enzyme phenotypes were also found in erythrocytes, platelets and cultured blood macrophages indicating that these cells have a common stem cell which is the site of the abnormality in CML. In the one studied patient, no evidence was found for involvement of cultured marrow fibroblasts. Clonal origin of CML virtually excludes cell recruitment as a sole pathogenetic mechanism. Either the leukemia arises as a consequence of a rare initial event in a single cell, or a series of events occurs in a clone such that it evolves into CML, or both.
... Attempts have also been made to assess clonal complexity during human hematopoiesis. Initial studies relied on X-chromosome inactivation patterns in hematological malignancies and were focused on the clonal origins of leukemia [88,89]. For instance, Fialkow et al . ...
... For instance, Fialkow et al . [89] assessed G6PD phenotypes in women to determine that chromic myelocytic leukemia CML was clonal in origin: single enzyme phenotypes were found in CML granulocytes but not in nonleukemic granulocytes. More recent studies have relied on hematopoietic output from patients receiving lentiviral gene therapy via HSCT [90,91]. ...
Hematopoietic stem and progenitor cells (HSPCs) govern the daily expansion and turnover of billions of specialized blood cells. Given their clinical utility, much effort has been made towards understanding the dynamics of hematopoietic production from this pool of stem cells. An understanding of HSC clonal dynamics during blood ontogeny could yield important insights into hematopoietic regulation, especially during aging and repeated exposure to hematopoietic stress; insults that may predispose individuals to the development of hematopoietic disease. Here, we review the current state of research regarding the clonal complexity of the hematopoietic system during embryogenesis, adulthood and hematologic disease.
... The technology that allowed this to be clarified arose from work by Philip Fialkow in hematological malignancies. Dr. Fialkow was a geneticist who showed that leukemias were clonal in women who were heterozygous for a variant in the X chromosome-linked G6PD gene, which resulted in G6PD proteins with distinct electrophoretic mobility (1). ...
Background: Very little was known about the molecular pathogenesis of thyroid cancer until the late 1980s. As part of the Centennial celebration of the American Thyroid Association, we review the historical discoveries that contributed to our current understanding of the genetic underpinnings of thyroid cancer. Summary: The pace of discovery was heavily dependent on scientific breakthroughs in nucleic acid sequencing technology, cancer biology, thyroid development, thyroid cell signaling, and growth regulation. Accordingly, we attempt to link the primary observations on thyroid cancer molecular genetics with the methodological and scientific advances that made them possible. Conclusions: The major genetic drivers of the common forms of thyroid cancer are now quite well established and contribute to a significant extent to how we diagnose and treat the disease. However, many challenges remain. Future work will need to unravel the complexity of thyroid cancer ecosystems, which is likely to be a major determinant of their biological behavior and on how they respond to therapy.
... Chronic myeloid leukemia (CML) is a myeloproliferative malignancy that arises from hematopoietic stem cells (1), and it accounts for 15% of adult leukemias (2). CML classically progresses through three phases (chronic phase, accelerated phase, and blast phase) and becomes more resistant to treatment in each successive phase (3). ...
The phenomenon of multidrug resistance (MDR) is called chemoresistance with respect to the treatment of cancer, and it continues to be a major challenge. The role of N-glycosylation in chemoresistance, however, remains poorly understood. Here, we established a traditional model for adriamycin resistance in K562 cells, which are also known as K562/ADR cells. Lectin blot, mass spectrometry, and RT-PCR analysis showed that the expression levels of N-acetylglucosaminyltransferase III (GnT-III) mRNA and its products, bisected N-glycans, are significantly decreased in K562/ADR cells, compared with the levels in parent K562 cells. By contrast, the expression levels of both P-glycoprotein (P-gp) and its intracellular key regulator, NF-κB signaling, are significantly increased in K562/ADR cells. These upregulations were sufficiently suppressed by the overexpression of GnT-III in K562/ADR cells. We found that the expression of GnT-III consistently decreased chemoresistance for doxorubicin and dasatinib, as well as activation of the NF-κB pathway by tumor necrosis factor α (TNFα), which binds to two structurally distinct glycoproteins, TNF receptor 1 (TNFR1) and TNF receptor 2 (TNFR2), on the cell surface. Interestingly, our immunoprecipitation analysis revealed that only TNFR2, but not TNFR1, contains bisected N-glycans. The lack of GnT-III strongly induced TNFR2's auto-trimerization without ligand stimulation, which was rescued by the overexpression of GnT-III in K562/ADR cells. Furthermore, the deficiency of TNFR2 suppressed P-gp expression while it increased GnT-III expression. Taken together, these results clearly show that GnT-III negatively regulates chemoresistance via the suppression of P-gp expression, which is regulated by the TNFR2-NF/κB signaling pathway.
... Preud'homme et al. (6) suggested that the T cells, as well as the conventional myeloma cells, were derived from myeloma stem cells by alternative differentiation within the same clone. It is relevant to point out that in human chronic granulocytic leukemia, alternative differentiation has been proposed to account for the observation that malignant myeloid cells and nonmalignant erythroid cells appear to be progeny of the same stem cell (26). However, in our studies with 315/+ we found that IBL were derived from the host and not from the tumor. ...
MOPC-315 is a BALB/c plasmacytoma that produces an IgA anti-TNP antibody (M315). 315/+ is a line derived from MOPC-315 and differentiates during in vivo growth from M315-producing, nonsecreting, lymphocytoid cells to large M315-secreting plasmacytoid cells. Studies were conducted to determine whether the small M315-bearing mononuclear cells, which account for more than one-third of the circulating mononuclear cells in mice with 315/+ tumors, were myeloma cells or host lymphocytes. Experiments conducted in CBF1 mice demonstrated that the circulating M315-bearing cells were of F1, rather than tumor, origin. Serologic and ultrastructural studies demonstrated that the cells were post-thymic T lymphocytes. After proteolytic removal, surface M315 was reexpressed in vitro by 315/+ cells, but not by T cells. M315-bearing T cells accounted for 0 to 4% of circulating mononuclear cells in mice with 315/P, a variant of MOPC-315 in which all cells synthesize M315, but only 2% of the cells are secretory. There was no obvious relationship between the frequency of M315-bearing lymphocytes and development of the humoral immunodeficiency that accompanies myeloma. These findings 1) identify an association between high levels of M315 secretion and development of M315-bearing T cells; 2) favor the view that M315 is acquired, rather than produced by host T cells; 3) raise the possibility that T cells with IgA-Fc receptors may be increased in mice with 315/+ tumors; and 4) are discussed relative to the mechanisms that have been proposed in the literature to account for the occurrence and significance of circulating, M-component-bearing lymphoid cells in murine and human myeloma.
... Blast crisis withstands only a few months and is identified by the myeloid population's expeditious growth and lymphoid differentiation apprehended blast cells (60). CML is consistently related to an acquired genetic aberration in the Philadelphia chromosome (Ph1), a shortened chromosome 22 resulting from reciprocal translocation of the long terms of chromosomes 9 and 22 (61,62). This translocation gives rise to the BCR/ABL fusion gene, which consequentially translates into the p210 BCR/ABL oncoprotein (63,64) in CML patients. ...
... Chronic myelogenous leukemia (CML) is an acquired clonal disease originating from pluripotent hematopoietic stem cells [230], and the inhibition of autophagy may increase therapeu-tic efficacy and improve patient prognosis [231]. Imatinib is a tyrosine kinase inhibitor widely used to treat CML and has a good therapeutic effect. ...
The Kelch-like (KLHL) family members consist of three domains: bric-a-brac, tramtrack, broad complex/poxvirus and zinc finger domain, BACK domain and Kelch domain, which combine and interact with Cullin3 to form an E3 ubiquitin ligase. Research has indicated that KLHL family members ubiquitinate target substrates to regulate physiological and pathological processes, including tumorigenesis and progression. KLHL19, a member of the KLHL family, is associated with tumorigenesis and drug resistance. However, the regulation and cross talks of other KLHL family members, which also play roles in cancer, are still unclear. Our review mainly explores studies concerning the roles of other KLHL family members in tumor-related regulation to provide novel insights into KLHL family members.
... CML is a clonal disease arising from a pluripotent HSC, as revealed by studies in the late 1960s of the expression of the glucose-6-phosphate dehydrogenase (G-6-PD) isoenzymes A and B within cells from three women with CML [7,8]. For normal cells, only one G-6-PD gene is active in each female cell, due to X chromosome inactivation, and they are a mixture of cells that express either the A or B isoenzyme. ...
There is compelling evidence to support the view that the cell-of-origin for chronic myeloid leukemia is a hematopoietic stem cell. Unlike normal hematopoietic stem cells, the progeny of the leukemia stem cells are predominantly neutrophils during the disease chronic phase and there is a mild anemia. The hallmark oncogene for chronic myeloid leukemia is the BCR-ABLp210 fusion gene. Various studies have excluded a role for BCR-ABLp210 expression in maintaining the population of leukemia stem cells. Studies of BCR-ABLp210 expression in embryonal stem cells that were differentiated into hematopoietic stem cells and of the expression in transgenic mice have revealed that BCR-ABLp210 is able to veer hematopoietic stem and progenitor cells towards a myeloid fate. For the transgenic mice, global changes to the epigenetic landscape were observed. In chronic myeloid leukemia, the ability of the leukemia stem cells to choose from the many fates that are available to normal hematopoietic stem cells appears to be deregulated by BCR-ABLp210 and changes to the epigenome are also important. Even so, we still do not have a precise picture as to why neutrophils are abundantly produced in chronic myeloid leukemia.
... Blast crisis withstands only a few months and is identified by the myeloid population's expeditious growth and lymphoid differentiation apprehended blast cells (60). CML is consistently related to an acquired genetic aberration in the Philadelphia chromosome (Ph1), a shortened chromosome 22 resulting from reciprocal translocation of the long terms of chromosomes 9 and 22 (61,62). This translocation gives rise to the BCR/ABL fusion gene, which consequentially translates into the p210 BCR/ABL oncoprotein (63,64) in CML patients. ...
Chronic myeloid leukemia (CML) is potentially fatal blood cancer, but there is an unmet need to discover novel molecular biomarkers. The hypothesis of this study aimed to elucidate the relationship of HIF1α with the redox system, Krebs cycles, notch1, and other regulatory proteins to better understand the pathophysiology and clinical relevance in chronic myeloid leukemia (CML) patients, as the molecular mechanism of this axis is still not clear. This study included CML patient samples (n = 60; 60: blood; 10: bone marrow tissues) and compared them with healthy controls (n = 20; blood). Clinical diagnosis confirmed on bone marrow aspiration, marrow trephine biopsy, and BCR/ABL1 translocation. Cases were subclassified into chronic, accelerated, and blast crises as per WHO guidelines. Molecular experiments included redox parameters, DNA fragmentation, Krebs cycle metabolites, and gene expression by RT-PCR/Western blot/LC-MS, PPI (STRING), Pearson correlation, and ROC curve analysis. Here, our findings show that p210/p190BCR/ABL1 translocation is common in all blast crisis phases of CML. Redox factor/Krebs oncometabolite concentrations were high, leading to upregulation and stabilization of HIF1α. HIF1α leads to the pathogenesis in CML cells by upregulating their downstream genes (Notch 2/4/Ikaros/SIRT1/Foxo-3a/p53, etc.). Whereas, downregulated ubiquitin proteasomal and apoptotic factors in CML pateints, can trigger degradation of HIF1α through proline hydroxylation. However, HIF1α showed a negative corelation with the notch1 pathway. Notch1 plays a tumor-suppressive role in CML and might have the potential to be used as a diagnostic marker along with other factors in CML patients. The outcome also revealed that oxidant treatment could not be effective in augmentation with conventional therapy because CML cells can enhance the levels of antioxidants for their survival. HIF1α might be a novel therapeutic target other than BCR/ABL1 translocation.
... Chronic myeloid leukaemia (CML) is a haematological malignancy that arises within a pluripotent haemopoietic stem cell. 1,2,3 The defining molecular aberration is a translocation involving the long arms of chromosome 9 and 22 (t [9;22][q34;q11.2]) which results in a chimeric breakpoint cluster region-Abelson murine leukaemia viral oncogene homolog 1 (BCR-ABL1) fusion gene encoding for a constitutively active tyrosine kinase. ...
Background: Chronic myeloid leukaemia (CML) is a haematological malignancy characterised by the translocation t(9;22)(q34;q11.2), resulting in a constitutively active tyrosine kinase. Globally, overall survival of blast crisis phase (BC) CML is one year. Newer tyrosine kinase inhibitors and allogeneic stem cell transplantation offer remission; however, refractory and relapsed disease remain the biggest challenges.
Objective: In South Africa, literature is lacking on BC-CML. This study aimed to determine the disease characteristics and overall survival in South Africa.
Methods: This retrospective, laboratory-based study reviewed all new BC-CML diagnoses via flow cytometry at Charlotte Maxeke Johannesburg Academic Hospital in Johannesburg, South Africa, between April 2016 and October 2019. BC-CML was defined as the presence of 20% blasts with a CML history or the BCR-ABL1 fusion gene (p210/p190) in the appropriate clinical or pathological context. Survival outcomes were inferred from clinical and laboratory data.
Results: Twenty-two new cases of BC-CML were diagnosed (median age: 34 years). There were 20 (91%) cases with the fusion transcripts p210 and two (9%) cases with p190 BCRABL1. For blast lineage, 14 cases were myeloid (63.6%), six were lymphoid (27.3%), and two were ambiguous (9.1%). There was a 72.7% mortality (16 cases); sepsis, refractory and relapsed disease were the major causes. Patients who achieved remission had lower blast percentages, simple karyotypes, and a trend towards higher white cell and platelet counts at presentation.
Conclusion: Optimised management of early-stage CML, prevention and aggressive management of sepsis, with advocation for newer therapies are needed to improve the overall survival of BC-CML in South Africa.
... The presence of CSCs has been assumed for several decades, as many tumors exhibit significant heterogeneity regarding their morphology, genetic lesions, cell proliferation kinetics and response to therapy even when the tumor initiated from a single cell [14]. The first possible relationship between cancer and stem cells was found in the hematopoietic system, when Fialkow and colleagues showed stem cell involvement in chronic myelocytic leukemia and acute non-lymphocytic leukemia [15,16]. Hierarchical tumor organization was also shown in solid cancers, as for instance only CD44+/CD24-/low tumorigenic breast cancer cells were able to form tumors that contained additional CD44+/CD24-/low cells, as well as phenotypically different non-tumorigenic cells [17]. ...
Endometrial cancer is one of the most common malignant diseases in women worldwide, with an incidence of 5.9%. Thus, it is the most frequent cancer of the female genital tract, with more than 34,000 women dying, in Europe and North America alone. Endometrial Cancer Stem Cells (CSC) might be drivers of carcinogenesis as well as metastatic and recurrent disease. Therefore, targeting CSCs is of high interest to improve prognosis of patients suffering of advanced or recurrent endometrial cancer. This review describes the current evidence of molecular mechanisms in endometrial CSCs with special emphasis on MYC and NF-κB signaling as well as mitochondrial metabolism. Furthermore, the current status of immunotherapy targeting PD-1 and PD-L1 in endometrial cancer cells and CSCs is elucidated. The outlined findings encourage novel therapies that target signaling pathways in endometrial CSCs as well as immunotherapy as a promising therapeutic approach in the treatment of endometrial cancer to impede cancer progression and prevent recurrence.
... La LMC es el resultado de la transformación neoplásica de una célula madre hematopoyética pluripotencial caracterizada inicialmente por hiperplasia mieloide, basofilía y esplenomegalia. (Fialkow et al, 1997). ...
En la era de la terapia con Imatinib mesilato, la evolución citogenética clonal sigue siendo asociada con supervivencia adversa en pacientes en fase crónica y acelerada, además la frecuencia en la aparición de evolución policlonal es más alta que con los tratamientos anteriores; y aunque su pronóstico es desconocido aún, la mayoría de estas anormalidades cromosómicas en células Ph- son típicas de los síndromes mielodisplásicos y leucemias mieloides agudas. Esta observación resalta la necesidad de monitorear a los pacientes LMC Ph+ que reciban Imatinib mesilato en donde además de buscar el transcrito BCR-ABL y el cromosoma Ph, se deberá realizar cariotipo con la finalidad de monitorear la aparición de alteraciones citogenéticas adicionales, aun y se responda adecuadamente a el tratamiento.
... CML is a type of myeloproliferative disorder caused by the malignant transformation of hematopoietic stem cells (Fialkow et al. 1977). On the molecular level, the outcome of the reciprocal translocation t (9;22) (q34q11) in CML is the formation of an oncogene termed BCR/ABL, which encodes a chimeric protein, BCR-ABL, with constitutive tyrosine kinase activity (Sun et al. 2013). ...
Imatinib (IM), targeting of BCR-ABL1 tyrosine kinase, is currently one of the first-line choices in the treatment of chronic myeloid leukemia (CML). This study aims to explore the molecular mechanisms underlying IM resistance in CML treatment. 108 CML patients were recruited and grouped according to their sensitivity to IM as the responder group (N = 66) and the non-responder group (N = 42). Real-time quantitative PCR (RT-qPCR) was performed to evaluate the expression of candidate circular RNAs (circRNAs), microRNA (miRNAs) and messenger RNA (mRNAs). No significant difference was noted regarding demographic and clinicopathological characteristics between the responder group and the non-responder group. The expression of circ_0080145, circ_0051886 and ABL1 mRNA was significantly increased, while the expression of miR-203 and miR-637 was decreased in the non-responder group as compared with the responders. By using in-silicon analysis, it was predicted that circ_0080145 and circ_0051886 targeted miR-203 and miR-637 respectively, and ABL1 was found to be shared direct target gene of miR-203 and miR-637. Ectopic over-expression of circ_0080145 and circ_0051886 respectively reduced the expression of miR-203 and miR-637. The expression of ABL1 mRNA/protein was most upregulated in culture cells co-transfected with circ_0080145 and circ_0051886 as compared with those cells individually transfected. This study established the signaling pathways of circ_0080145/miR-203/ABL1 and circ 0051886/miR-637/ABL1. The deregulation of circ_0080145 and circ_0051886 is, at least partially, responsible for the development of IM chemoresistance in CML by regulating expression of ABL1 via modulating expression of miR-203 and miR-637.
... They arise in a tissue-specific stem or progenitor cell that is able to give rise to the different types of functional cells within a tissue. It is well-known that chronic myeloid leukemia (CML) originates in a hematopoietic stem cell (HSC) [2], but paradoxically the offspring of the leukemia stem cells (LSCs) are restricted to development just along the granulocyte pathway. Similarly, acute erythroid leukemia is also an HSC disorder, and the leukemia cells belong to the erythroid lineage only. ...
In principle, an oncogene is a cellular gene (proto-oncogene) that is dysfunctional, due to mutation and fusion with another gene or overexpression. Generally, oncogenes are viewed as deregulating cell proliferation or suppressing apoptosis in driving cancer. The cancer stem cell theory states that most, if not all, cancers are a hierarchy of cells that arises from a transformed tissue-specific stem cell. These normal counterparts generate various cell types of a tissue, which adds a new dimension to how oncogenes might lead to the anarchic behavior of cancer cells. It is that stem cells, such as hematopoietic stem cells, replenish mature cell types to meet the demands of an organism. Some oncogenes appear to deregulate this homeostatic process by restricting leukemia stem cells to a single cell lineage. This review examines whether cancer is a legacy of stem cells that lose their inherent versatility, the extent that proto-oncogenes play a role in cell lineage determination, and the role that epigenetic events play in regulating cell fate and tumorigenesis.
... Chronic myeloid leukemia (CML) is an ideal disease to explore non-mutational mechanisms of drug-insensitivity. It is a clonal disorder originating from hematopoietic stem cells (HSCs) due to a single oncogenic fusion protein BCR-ABL, a constitutively active tyrosine kinase [2,3]. The standard molecular targeted therapy for CML is tyrosine kinase inhibitors (TKI), which eradicates the bulk of the disease but not the quiescent LSCs [4][5][6][7][8]. ...
Leukemic stem cells (LSCs) can acquire non-mutational resistance following drug treatment leading to therapeutic failure and relapse. However, oncogene-independent mechanisms of drug persistence in LSCs are incompletely understood, which is the primary focus of this study. We integrated proteomics, transcriptomics, and metabolomics to determine the contribution of STAT3 in promoting metabolic changes in tyrosine kinase inhibitor (TKI) persistent chronic myeloid leukemia (CML) cells. Proteomic and transcriptional differences in TKI persistent CML cells revealed BCR-ABL-independent STAT3 activation in these cells. While knockout of STAT3 inhibited the CML cells from developing drug-persistence, inhibition of STAT3 using a small molecule inhibitor sensitized the persistent CML cells to TKI treatment. Interestingly, given the role of phosphorylated STAT3 as a transcription factor, it localized uniquely to genes regulating metabolic pathways in the TKI-persistent CML stem and progenitor cells. Subsequently, we observed that STAT3 dysregulated mitochondrial metabolism forcing the TKI-persistent CML cells to depend on glycolysis, unlike TKI-sensitive CML cells, which are more reliant on oxidative phosphorylation. Finally, targeting pyruvate kinase M2, a rate-limiting glycolytic enzyme, specifically eradicated the TKI-persistent CML cells. By exploring the role of STAT3 in altering metabolism, we provide critical insight into identifying potential therapeutic targets for eliminating TKI-persistent LSCs.
... CSC hypothesis provides evidence that relatively sparse cell population within a tumor possess common measures with somatic stem cells representing the capacity for infinite self-renewal and initiate to drive the tumor growth (Huntly & Gilliland, 2005). Several evidences for putative CSCs have been described in blood (Fialkow et al., 1977), breast (Al-Hajj et al., 2003), lungs (Eramo et al., 2008), prostate (Collins et al., 2005), colon (Bussolati et al., 2011), liver (Chiba et al., 2007), pancreas , and brain (Singh et al., 2004). If we target CSCs or their immediate progeny, then cancer risk could gradually be decreased. ...
Cancer, being the leading cause of death in the globe, has been one of the major thrust areas of research worldwide. In a new paradigm about neoplastic transformations, the initiation and recurrence of disease is attributed to few mutated cells in bulk of tumor called cancer stem cells (CSCs). CSCs have capacity of self-renewal and differentiation, which are known for resistance to radio and chemotherapy leading to recurrence of the disease even after treatment. Most of traditional drugs implicated in cancer therapy targeting primary tumors have substantial toxicity to the physiological system and have not been efficient in targeting these CSCs leading to poor prognosis. Targeting these CSCs in bulk of tumor might be novel strategy for cancer chemoprevention and therapeutics. Diet-derived interventions and diverse natural products are known to target these CSCs and related signaling pathways, namely, Wnt, Notch, and Hedgehog pathways, which are implicated for CSC self-renewal.
Practical applications
Cancer remains a global challenge even in this century. Poor prognosis, survival rate, and recurrence of the disease have been the major concerns in traditional cancer therapy regimes. Targeting cancer stem cells might be novel strategy for elimination and cure of the chronic disease as they are known to modulate all stages of carcinogenesis and responsible for recurrence and resistance to chemotherapy and radiotherapy. The evidence support that natural products might inhibit, delay, or reverse the process of tumorigenesis and modulate the different signaling pathways implicated for cancer stem cells self-renewal and differentiation. Natural products have minimal toxicity compared to traditional cancer therapy drugs since they have long been utilized in our food habits without any major side effects reported. Thus, targeting cancer stem cells with natural product might be a novel strategy for drug development in cancer chemoprevention and therapeutics.
... Studies using various markers have shown that CML is a clonal neoplasm that involves a multipotent progenitor cell capable of lymphoid as well as myeloid differentiation. [1][2][3] The genes for glucose-6-phosphate dehydrogenase (G6PD), phosphoglycerate kinase, hypoxanthine phosphoribosyltransferase (HPRT), M27 and human androgen receptor are located on the X chromosome, which undergoes random inactivation during early embryogenesis in each somatic cell in females, and thus the DNAs or their products have been used for the clonal analysis of CML. 4 Clonal analysis using these genes or their products, however, is limited to samples only with a homogenous leukemia cell population. On In a unique case of CML arising as a clone in the monosomic (45, Xc) cells in a 27-year-old patient with Turner syndrome mosaic, extensive cytogenetic studies at diagnosis, including triple color fluorescence in situ hybridization (FISH), showed that Ph-positive monosomic leukemia cells predominated, with a few Ph-negative monosomic and normal diploid (46, XX) cells. ...
In a 27-year-old female with Turner syndrome mosaic, Philadelphia (Ph) chromosome-positive chronic myeloid leukemia (CML) occurred only in the monosomic cells (45, Xc). Extensive cytogenetic studies, including triple-color fluorescence in situ hybridization (FISH), revealed that Ph-positive monosomic cells (45, Xc), Ph-negative monosomic cells and normal diploid cells (46, XX) were present in her bone marrow at diagnosis. After successful interferon therapy, the non-leukemia cells expanded and reconstituted normal hematopoiesis resulting in complete cytogenetic response, following the selective suppression of the monosomic Ph-positive leukemia clone. The ratio of Xc to XX cells in bone marrow cells was significantly increased to that in skin fibroblasts. Moreover, the ratio of Ph-positive cells to Ph-negative cells was found to be significantly different between karyotyping and FISH. Studies of this quite unique case not only confirmed the clonality of CML, effectiveness of interferon-α and X chromosome imbalance among different tissues, but also demonstrated a discrepant increase of the BCR/ABL-positive clone in CML. The latter supports the hypothesis that reduced programmed cell death may be the primary mechanism responsible for the expansion of the leukemia clone in CML. Our study verifies the importance of extensive analysis of a neoplastic disease in patients with a constitutional chromosomal abnormality.
... This condition is observed in myelodysplastic syndromes (MDS), 1-3 paroxysmal nocturnal hemoglobinuria (PNH), 4 and myeloproliferative disorders (MPD) 5,6 including chronic myeloid leukemia (CML). 7,8 The clonal nature of these hematological disorders has been analyzed using mainly peripheral blood samples. To understand the mechanism underlying clonal hematopoiesis, however, investigators need to know the clonality of hematopoietic progenitor cells in bone marrow as well as peripheral blood cells. ...
X chromosome inactivation and polymorphism of the human androgen receptor (HUMARA) gene has been applied for analyzing the clonality of blood cells. In the present study, the clonal relationship was investigated between peripheral blood polymorphonuclear cells (PMNCs) and marrow progenitor cells and the origin of ringed sideroblasts in patients with refractory anemia with ring sideroblasts (RARS) by polymerase chain reaction (PCR) of HUMARA gene. The X-inactivation patterns of circulating PMNCs and T lymphocytes as well as individual granulocyte colonies grown in vitro from bone marrow cells were analyzed. The development of ringed sideroblasts in erythroid colonies by iron staining and their X-inactivation pattern were also examined. All three RARS patients showed monoclonal PMNCs. In granulocyte colonies, however, two different X-inactivation patterns were observed in all patients, indicating that non-clonal progenitor cells remained in the bone marrow. All erythroid colonies consisted of ringed sideroblasts exclusively showed one pattern dominant in those of PMNCs. Our findings suggest that non-clonal progenitor cells persist in some RARS cases, that erythroid progenitors show mosaicism, and that ringed sideroblasts may be derived from an abnormal clone involved in the pathogenesis of this disease.
... However, not every HSC barcode can be found amongst CD4 T cells or CD8 T cells in the peripheral blood ( Fig. 1D-E, and S1), suggesting that some engrafted HSC clones do not contribute to the mature T cell repertoire. This explains why a chronic myelocytic leukemia clone from the HSC pool can be found in granulocytes, monocytes, erythrocytes, and B cells, but is rarely present in T cells (41). It is likely that the migration of the T cell precursors, mainly the common lymphocyte progenitors (CLP), to the thymus or the maturation of T cells within the thymus is episodic and restricts the number of HSC clones that eventually contribute to mature T cells (42). ...
While hematopoietic stem cells (HSCs) have been extensively studied at the population level, little is known about the lineage commitment of individual clones. Here, we provide comprehensive maps of in vivo HSC clonal development in mice under homeostasis and after depletion of the endogenous hematopoietic system. Under homeostasis, all donor-derived HSC clones regenerate blood homogeneously throughout all measured stages and lineages of hematopoiesis. In contrast, after the hematopoietic system has been depleted by irradiation or by an anti-ckit antibody, only a small fraction of donor-derived HSC clones differentiates while dominantly expanding and exhibiting lineage bias. We identified the cellular origins of clonal dominance and lineage bias, and uncovered the lineage commitment pathways that lead HSC clones to differential blood production. This study reveals surprising alterations in HSC regulation by irradiation, and identifies the key hematopoiesis stages that may be manipulated to control blood production and balance.
SIGNIFICANCE STATEMENT
Hematopoietic stem cells (HSCs) sustain daily blood production through a complex step-wise lineage commitment process. In this work, we present the first comprehensive study of HSC lineage commitment at the clonal level and identify new HSC regulatory mechanisms that are undetectable by conventional population level studies. First, we uncover distinct HSC clonal pathways that lead to differential blood production and imbalances. Second, we reveal that HSC regulation under physiological conditions is strikingly different from that after injury. Third, we present a comprehensive map of HSC activities in vivo at the clonal level.
... Finally, the epigenetic priming model supports the idea of a plastic, easily reprogrammable, stem cell as a leukemia-initiating cell, a property that may be used to target the LSC. l In the case of leukemias, the stem nature of the LIC was clear many years ago for chronic myeloid leukemia (CML, [38]) and was the first to be experimentally demonstrated in acute myeloid leukemia (AML [18,39]), followed by Philadelphia-positive acute lymphoblastic leukemia (ALL) [19] and, more recently, also T-ALLs [23,40]. This is supported by the fact that animal models of different leukemias generated by targeting the cancerinitiating lesions to mouse hematopoietic stem and progenitor cells (HSPCs) are capable of recapitulating the respective malignancies associated in humans with the lesion used (see below, Subheading 5.2.2) [27,28,30]. ...
Only 10 years ago, the existence of cancer stem cells (CSCs) was still hotly debated. Even today, when their presence in most tumor types has been clearly demonstrated, all the consequences of their existence are far from being realized neither in the clinic nor, very often, in basic and translational cancer research. The existence of CSCs supposes a true change of paradigm in our understanding of cancer, but it will only have a real impact when we will properly assimilate its implications and apply these insights to both cancer research and cancer treatment. In this primer to the topic of leukemia stem cells (LSCs) our aim is to highlight with broad brushstrokes the most relevant of their properties, how these characteristics led to their identification, and the implications that the existence of LSCs has for the research and fight against leukemia.
... Von allen Krebsentitäten im Kindesalter repräsentieren Leukämien ungefähr 30 % (Kaatsch et al., 2016). Die chronisch myeloische Leukämie (CML) gehört zu den myeloproliferativen Erkrankungen und ist mit einer Inzidenz von 0,06 -0,12 pro 100 000 Kinder pro Jahr im Vergleich zu 0,7-1,8 pro 100 000 Erwachsene pro Jahr sehr selten (de la Fuente et al., 2014;Fialkow et al., 1977;Smith et al., 2014;Chen et al., 2012). Im Median wird die CML bei Kindern im Alter von 12,4 Jahren diagnostiziert, wobei das männliche Geschlecht etwas häufiger (1,2 -1, 5×) als das weibliche betroffen ist (Millot et al., 2005;Millot et al., 2014c). ...
Hintergrund und Ziele: Die chronisch myeloische Leukämie (CML) repräsentiert eine seltene Leukämieform im Kindesalter. Das ursächliche Fusionsgen aus den Genen Breakpoint cluster region (BCR) und Abelson murine leukemia (ABL) resultiert in einer ständig aktiven Tyrosinkinase. Dies führt zu einer Fehlregulation myeloischer Stammzellen mit einer vermehrten Zellteilung. Trotz moderner Therapiemethoden mit Tyrosinkinaseinhibitoren (TKIs) sprechen manche Patienten nicht ausreichend auf die Erstlinientherapie mit Imatinib, einem TKI der ersten Generation, an. Bestimmte Varianten des menschlichen Genoms, sogenannte Polymorphismen, wurden mit einem verzögerten Therapieerfolg bei erwachsenen Patienten in Zusammenhang gebracht. Sie bieten im Allgemeinen die Möglichkeit, mit dem Auftreten einer Erkrankung in Verbindung zu stehen und als prognostische Marker für das Therapieansprechen zu fungieren. Daten, die den Einfluss der Polymorphismen auf die kindliche CML beschreiben, fehlen bislang. Ziel war es, Kinder mit CML auf ausgewählte Polymorphismen zu untersuchen und deren Auswirkung auf den Therapieerfolg mit TKIs zu eruieren. Methoden: In der vorliegenden Dissertation wurden Knochenmarks- und lutproben von 101 Kindern mit BCR-ABL-positiver CML untersucht. Die Gene B-cell chronic lymphocytic leukemia-lymphoma like 11 gene (BIM), ATP binding cassette subfamily B member 1 (ABCB1) und B-cell leukemia/lymphoma 2 (BCL2) waren dabei Gegenstand der Analysen. Zum Zeitpunkt der Probenentnahme befanden sich die Patienten in klinischer Remission. Die Genotypisierung der Polymorphismen erfolgte mithilfe der Polymerase-Kettenreaktion (PCR), Restriktion, Agarose-Gelelektrophorese und einer extern durchgeführten Sequenzierung nach Sanger. Im zweiten Teil dieser Arbeit wurden die genomischen Varianten aus einer Whole Exome Sequencing (WES)-Analyse mittels PCR und externer Sequenzierung validiert. Untersucht wurden Initial- und Remissionsproben von 33 pädiatrischen CML-Patienten aus Blut oder Knochenmark. Die Vergleichspopulation bestand aus 49 adulten CML-Patienten. Abschließend wurden die genomischen Varianten in somatische Mutationen und Keimbahnmutationen gegliedert. Ergebnisse und Beobachtungen: Zwischen dem Auftreten einer Erkrankung und dem Genotyp der untersuchten Polymorphismen auf dem BIM-Gen wurde auf dem 5%-Signifikanzniveau kein Zusammenhang gefunden. Entgegen der Erwartungen waren im Bezug auf die Einzelnukleotid-Polymorphismen (SNPs)rs2032582 (ABCB1) und rs1801018 (BCL2) unter den pädiatrischen CML-Patienten die Genotypen T/T beziehungsweise A/A häufiger vertreten als in einer europäischen Vergleichsgruppe gesunder Erwachsener (T/T vs. G/G-G/T-A/G-A/T, p = 0, 02; A/A vs. G/G-A/G, p = 0, 04).Der Genotyp G/T (ABCB1, rs2032582) wurde dagegen bei den erkrankten Kindern seltener festgestellt als in der gesunden Erwachsenenpopulation (p = 0, 02). Die Vermutung, die Polymorphismen seien mit einer erhöhten initialen Leukozytenzahl im Blut bei Diagnosestellung der CML assoziiert, konnte nicht bestätigt werden. Die in einer chinesischen Population beschriebene BIM-Deletion wurde im pädiatrischen Patientenkollektiv in Übereinstimmung mit einer deutschen Referenzpopulation nicht nachgewiesen. Denkbar ist eine ethnische Abhängigkeit des BIM-Deletion-Polymorphismus. Die SNPs rs1045642, rs1128503, rs2032582 auf dem ABCB1-Gen und rs1801018 auf dem BCL2-Gen standen hingegen in einem signifikanten Zusammenhang mit dem Therapieansprechen. Patienten mit dem jeweiligen assoziierten Allel sprachen deutlich verzögert auf eine Therapie mit TKIs an (p < 0, 05). Besonders wenige Kinder mit einem C/C oder C/T-Genotyp an der Position des SNP rs1128503 erreichten eine molekulare Remission (BCR-ABL/ABL ≤ 0,1%) nach 18 Monaten TKI-Therapie im Vergleich zu jenen ohne C-Allel (p < 0, 001). Die Polymorphismen rs724710 auf dem BIM-Gen und rs2279115 auf dem BCL2-Gen schienen das Therapieansprechen der CML-Patienten nicht relevant zu beeinflussen (p > 0, 05). SNPs, die auf einem Gen liegen (ABCB1, BCL2), waren zudem nicht unabhängig voneinander (p < 0, 0001). Das zeigten Korrelationen der einzelnen SNPs untereinander. Dabei war der Zusammenhang zwischen dem Auftreten der ABCB1-SNPs rs1128503 und rs2032582 besonders stark ausgeprägt (Korrelationskoeffizient ϕn = 1). Schlussfolgerung: Zusammenfassend ließ sich im Falle des ABCB1 und BCL2-Gens ein unterschiedliches Ansprechen der pädiatrischen CML-Patienten auf eine Therapie mit TKIs abhängig vom zugrundeliegenden Genotyp des jeweiligen Polymorphismus beobachten. Besonders der SNP rs1128503 (ABCB1) war mit einer unzureichenden molekularen Antwort auf eine TKI-Therapie assoziiert. Im klinischen Alltag könnten Polymorphismen als relevante Orientierungshilfe bei der Auswahl geeigneter Therapeutika dienen. Gerade vor dem Hintergrund einer lebenslangen TKI-Therapie ab dem Kindesalter wäre ein prädiktiver Marker für das erfolgreiche Ansprechen und Aussetzen der Therapie in Phasen klinischer Remission von besonderem Interesse.
... Chronic myeloid leukaemia (CML) originates from the malignant transformation of a single pluripotent haematopoietic stem cell that confers the CML clone proliferative advantage. 1 Common symptoms of CML are secondary to that of anaemia and splenomegaly, including fatigue, weight loss, early satiety and left upper abdominal bloating. However, some asymptomatic CML patients are diagnosed incidentally after a routine blood test. 2 The introduction of tyrosine kinase inhibitors (TKIs), which target the molecular defect at BCR-ABL1 tyrosine kinase, 3 has transformed the outlook of CML treatment. ...
Purpose
Chronic myeloid leukaemia (CML) patients on long-term tyrosine kinase inhibitor (TKI) therapy are susceptible to drug-related problems (DRPs). This study aimed to evaluate the acceptability and outcomes of pharmacist-led interventions on DRPs encountered by CML patients.
Methods
This study included participants from the intervention arm of a randomised controlled trial which was conducted to evaluate the effects of pharmacist-led interventions on CML patients treated with TKIs. Participants were recruited and followed up in the haematology clinics of two hospitals in Malaysia from March 2017 to January 2019. A pharmacist identified DRPs and helped to resolve them. Patients were followed-up for six months, and their DRPs were assessed based on the Pharmaceutical Care Network Europe Classification for DRP v7.0. The identified DRPs, the pharmacist’s interventions, and the acceptance and outcomes of the interventions were recorded. A Poisson multivariable regression model was used to analyse factors associated with the number of identified DRPs per participant.
Results
A total of 198 DRPs were identified from 65 CML patients. The median number of DRPs per participants was 3 (interquartile range: 2, 4). Most participants (97%) had at least one DRP, which included adverse drug events (45.5%), treatment ineffectiveness (31.5%) and patients’ treatment concerns or dissatisfaction (23%). The 228 causes of DRPs identified comprised the following: lack of disease or treatment information, or outcome monitoring (47.8%), inappropriate drug use processes (23.2%), inappropriate patient behaviour (19.9%), suboptimal drug selection (6.1%), suboptimal dose selection (2.6%) and logistic issues in dispensing (0.4%). The number of concomitant medications was significantly associated with the number of DRPs (adjusted Odds Ratio: 1.100; 95% CI: 1.005, 1.205; p = 0.040). Overall, 233 interventions were made. These included providing patient education on disease states or TKI-related side effects (75.1%) and recommending appropriate instructions for taking medications (7.7%). Of the 233 interventions, 94.4% were accepted and 83.7% were implemented by the prescriber or patient. A total of 154 DRPs (77.3%) were resolved.
Conclusions
The pharmacist-led interventions among CML patients managed to identify various DRPs, were well accepted by both TKI prescribers and patients, and had a high success rate of resolving the DRPs.
... 3,4 Understanding the molecular basis of CML has allowed the development of targeted drugs against cancer cells. 5 Imatinib mesylate (IM), as a first-generation tyrosine kinase inhibitor (TKI), is used for the treatment of all newly diagnosed patients with CML. Once IM reaches the target cancer cells, the drug works by inhibiting the tyrosine kinase activity of the Bcr-Abl1 fusion protein. ...
Objective:
Tyrosine kinase inhibitors (TKIs) are considered standard first-line treatment in patients with chronic myeloid leukemia. Because ABL kinase domain mutations are the most common causes of treatment resistance, their prevalence and assessment during treatment may predict subsequent response to therapy.
Methods:
The molecular response in Bcr-Abl1IS was tested via quantitative real-time polymerase chain reaction. We used the direct sequencing technique to discover the mutations in the ABL kinase domain. The IRIS trial established a standard baseline for measurement - (100% BCR-ABL1 on the 'international scale') and a major molecular response (good response to therapy) was defined as a 3-log reduction in the amount of BCR-ABL1 - 0.1% BCR-ABL1 on the international scale.
Results:
We observed 11 different mutations in 13 patients, including E255K, which had the highest mutation rate. A lack of hematologic response was found in 22 patients, who showed a significantly higher incidence of mutations.
Conclusion:
Detection of kinase domain mutations is a reliable method for choosing the best treatment strategy based on patients' conditions, avoiding ineffective treatments, and running high-cost protocols in patients with acquired resistance to TKIs.
... Chronic myeloid leukemia is the uncontrolled proliferation and expansion of myeloid cells transformed from the early progenitor cells that include megakaryocytic cells, B-lymphoid cells, erythrocytes and monocytes [Fialkow et al., 1977]. Chronic myeloid leukemia is the first disease through which the specific genetic abnormality of Philadelphia chromosome in leukemia cells could b e linked to the events of pathogenetic during the initiation of leukemia [ Nowell and Hungerford, 1960]. ...
Wedelia trilobata belonging to the Asteraceae family contains the bio-active molecules especially molecule extracted through methanol showed the prominent antileukemia activity in the previous studies that buoyant not only for purification and structural analysis but also for the elucidation and confirmation of biological activities for specified phytoconstituent. Structural analysis studies on the extracted compound revealed the methoxy, methylphenyl and chromone groups containing molecule which is unique in its structure. The phytoconstituent is annealed with prominent anti-proliferation activity on chronic myeloid leukemia cells as told by Thymidine uptake and Wound healing assays. The antiproliferation is due to the induction of apoptosis which is confirmed by the DNA fragmentation and Flow cytometry assays. As per the pharmacological point of view the phytoconstituent obtained from our studies has unveiled the promising results against chronic myeloid leukemia.
... CML was first identified as a clonal hematological malignancy from pluripotent bone marrow stem cells [23,24]. Such finding increased our understanding of the stages of disease mechanism and helps in the diagnosis, differential diagnosis, and treatment. ...
Many human tumors originate from tissues that, as part of their normal physiology, undergo a process of constant turnover, sustained by minority populations of long-lived cells, called stem cells. In many tissues, stem cells are the only cells that can persist throughout the lifetime of the host, and therefore represent ideal targets for malignant transformation, because they can accumulate carcinogenic insults, such as genetic and epigenetic mutations, over long periods of time. Indeed, experimental evidence indicates that many forms of human cancer can be best understood as “aberrant organs” sustained in their growth and metastatic dissemination by a pathologic population of stemlike cells, originated as a result of mutations in genes that regulate key properties of the stem cell phenotype, such as the capacity for unlimited numerical expansion (self-renewal), the capacity to give rise to progenies of specialized cells (differentiation) and the capacity to migrate into new tissue locations and support tissue regeneration (tissue repair). This hypothesis is supported by the observation that, in many forms of human cancer, only a specific, phenotypically distinct, subset of cancer cells can form tumors when serially transplanted into immune-deficient mice (cancer stem cells), and implies that, to achieve cure, therapies must eradicate such subset of cancer cells. Studies on the gene-expression profile of cancer stem cell populations have led to the discovery of new prognostic biomarkers and pharmacological targets in cancer biology, and to the identification of tumor subtypes with differential response to anti-tumor drugs, opening new research avenues in clinical oncology.
Kronik miyeloid lösemi (KML), kemik iliğindeki hematopoietik kök hücrelerin monoklonal çoğalmasıyla karakterize myeloproliferatif bir hastalıktır. Gebelikte KML oldukça nadir görülür, tahmin edilen yıllık insidansı 100.000 gebelikte 1’dir. KML gebelikte görülen lösemilerin %10’undan azını oluşturur. Gebelikte löseminin tedavi ve bakım süreci, anne ve bebek sağlığı açısından son derece önemlidir. Lösemili gebe olguların hemşirelik bakımının önemine dikkat çekmek ve daha etkin bir hemşirelik bakımına ilişkin farkındalık oluşturmak amacı ile 27 yaşında, 5 aylık gebe, KML tanılı olgu ele alındı. KML tedavisi altında sağlıklı canlı doğum yaptırıldı. Gebelik ve pospartum hemşirelik bakımı gerçekleştirildi.
El objetivo principal de este estudio fue determinar la frecuencia de los transcritos de fusión de BCR-ABL por medio de la técnica de reacción en cadena de la polimerasa de transcripción reversa (RT-PCR) en muestras de médula ósea procedentes de pacientes con LMC y LLA-B para ayudar a la caracterización genética de la población leucémica guatemalteca. La relación entre la frecuencia de los transcritos del gen quimérico BCR-ABL y el tipo de leucemia asociada a sus características demográficas establece una relación significativa en base al grupo etario el cual presenta una tendencia poblacional que muestra dos picos de incidencia, los cuales corresponden al rango de 30-40 años en la expresión de b2a2 o b3a2 en pacientes con LMC y al de 5-10 años en la expresión de e1a2 en pacientes con LLA-B (P=0.022). La relación entre la frecuencia de los transcritos del gen quimérico BCR-ABL y las características clínicas demuestra una diferencia significativa respecto a la distribución del recuento plaquetario según el tipo de transcrito patológico expresado (P= 0.012). La expresión del transcrito e1a2 se encuentra relacionada a una actividad trombopoyética apreciablemente menor, mientras que el incremento del recuento plaquetario corresponde a la detección del transcrito b2a2. Estudios efectuados en diferentes regiones geográficas establecen diferencias significativas entre la frecuencia de expresión de los reordenamientos del gen quimérico BCR-ABL. En el presente estudio se determinó que la frecuencia de expresión de los distintos transcritos quiméricos del gen BCR-ABL en 35 muestras de médula ósea procedentes de pacientes guatemaltecos con LMC y LLA-B corresponde a una baja frecuencia para el transcrito e1a2 (11%) con respecto a la frecuencia de los transcritos b2a2 (43%) y b3a2 (43%) y con una frecuencia de coexpresión del 3% representada por los transcritos b2a2/b3a2. Pudo identificarse la predominancia del transcrito e1a2 en pacientes con LLA-B y de los transcritos b2a2 y b3a2, así como su coexpresión (b2a2/b3a2) en pacientes con LMC, sin embargo no se encontraron diferencias significativas en la frecuencia de expresión del transcrito b2a2 comparado con b3a2 (p=0.369). Se encontró que los hallazgos obtenidos de estudios efectuados en diferentes regiones geográficas del mundo en pacientes con LLA-B coinciden con la frecuencia reportada en la población leucémica guatemalteca con esta patología detectándose en el 100% de pacientes con LLA-B la expresión del transcrito patológico e1a2. Se puede concluir que la frecuencia de los transcritos de BCR-ABL entre la población guatemalteca con LMC concuerda con la reportada en estudios efectuados en países latinoamericanos, donde la población pertenece a un grupo mayoritariamente mestizo y es inversa a la reportada en países occidentales y orientales en donde existe predominio de poblacionescaucásicas y asiáticas, debido a que en dichos estudios se reporta frecuentemente una diferencia significativa entre la frecuencia de expresión de b2a2 (30-40%) comparado con la expresión de b3a2 (50-60%). Esto sugiere que la población leucémica guatemalteca que expresa los transcritos de fusión del gen quimérico BCR-ABL presenta una conducta biológica diferente respecto a la reportada en la población asiática y caucásica producto de la variabilidad genética entre poblaciones como posible explicación para las diferencias observadas en este estudio.
The latest version of the World Health Organization guidelines focuses mainly on the genetic and cytogenetic features of hematologic neoplasms as predictors of diagnostic, treatment decision, prognostic outcome, and for treatment monitoring in hematological malignancies. There are different techniques to identify these abnormalities. Live cells are needed for chromosome preparation. The Hematological malignancies include myeloid and lymphoid neoplasms. The myeloid neoplasms include Myelodysplastic syndromes, myeloproliferative neoplasms, and acute myeloid leukemias. The Lymphoid neoplasms include acute and chronic lymphocytic leukemias, plasma cell neoplasms, myeloma, hodgkin, and non-hodgkin lymphomas. The first chromosomal abnormality discovered in connection with cancer is the Philadelphia chromosome, which is an abnormal chromosome 22, formed due to the translocation between chromosomes 9 and 22. The presence of this abnormal chromosome confirms the diagnosis of “CML”. After that, hundreds of chromosomal abnormalities have been identified in hematological malignancies in different parts of the world. In AML, specific abnormalities were identified as having a good prognosis, intermediate prognosis, and poor prognosis. In other hematological malignancies also there some specific chromosome abnormalities are associated with prognostication. Now a day’s clinicians depend mainly on genetic abnormalities for the proper treatment management of hematological malignancies, so the study of chromosomal abnormalities is essential.
Introduction:
P-Loop mutations in CML patients prevent the conformational change in BCR-ABL1 necessary for drug binding. The present study aimed to evaluate the impact of mutations in this domain on the prognosis of the disease and also to associate the baseline Sokal relative risk score with the overall survival in non-responding CML patients.
Methods:
Blood samples were analyzed using ARMS-PCR and then an association was assessed between presence/absence of mutations, hematological and molecular response, disease progression, overall survival, and Sokal score.
Results:
Of the total 250 CML patients, 102 were found to be treatment-resistant. Fifty-three patients harbored P-Loop mutations with G250E (12.7%) being most frequent. Complete hematological response and major molecular response were achieved by only 27.7% and 5.7 patients, respectively. Worst survival (57.1%) was observed in Y253H positive patients while according to Sokal score in high-risk patients harboring Y253F (50%) and E255V (50%).
Conclusion:
The presence of P-Loop domain mutations negatively impacted the prognosis of the disease in terms of disease advancement and overall survival. So, the timely performance of the BCR-ABL1 mutational analysis and the modifications in the treatment plan based on the mutation identified would help in a better outcome of the disease.
I define the term as a combined concept for both tumor and leukemia. Tumor represents mass or solid tumor in a strict sense and leukemia seldom forms mass. However, in order for tumor to achieve hematogenous metastasis, the tumor cells need to make an entry into circulation as leukemic cells usually do. Some researchers call circulating tumor cells (CTC) . Both malignancy and neoplasm are synonymous with cancer. Here I describe unique features of cancer.
Although tyrosine kinase inhibitors (TKIs) play a crucial role in the treatment of chronic myeloid leukemia (CML) and Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph + ALL), intolerance and resistance to TKIs have been serious problems. Due to a lack of research, the importance of the pharmacokinetics (PK) of TKIs is currently unclear. We examined the PK of the third-generation TKI ponatinib to monitor side effects and efficacy during treatments for one patient with CML-chronic phase (CP-CML) and two who received allogeneic hematopoietic stem cell transplantation (allo-HSCT), one for CML-blastic crisis (BC-CML) and one for Ph + ALL. The patient with CP-CML was intolerant to multiple TKIs (dasatinib, nilotinib, imatinib, and bosutinib) and thus was switched to ponatinib (15 mg/day). The patients who received allo-HSCT for BC-CML and Ph + ALL received ponatinib (15 mg/day) as maintenance therapy. Notably, serial evaluation of the PK of ponatinib showed that the median trough values (ng/ml) were 17.2 (12.2–34.5), 33.1 (21.2–40.3) and 27.7 (13.6–29.9) in patients 1, 2, and 3, respectively. These values were around the target concentration (23 ng/ml). All patients are maintaining complete remission without side effects. In conclusion, serial evaluation of PK of ponatinib may yield meaningful information about its safety and efficacy.
Chronic myelogenous leukemia (CML) is characterized by a translocation involving the c-abl protein-tyrosine kinase gene. A chimeric mRNA is formed containing sequences from a chromosome 22 gene (bcr) at its 5' end and all but the variable exon 1 of c-abl sequence. The product of this mRNA, p210bcr-abl, has constitutively high protein-tyrosine kinase activity. We examined K562 cells and other lines established from CML patients for the presence of phosphotyrosine (P-Tyr)-containing proteins which might be p210bcr-abl substrates. Two-dimensional gel separation of 32P-labeled proteins followed by phosphoamino acid analysis of 25 phosphoproteins, which comprised the major alkali-stable phosphoproteins, indicated that three related proteins of 41 kDa are the most prominent P-Tyr-containing proteins detected by this method. The 41-kDa phosphoproteins are found in two other CML lines that we examined but not in lines of similar lineage isolated from patients with distinct leukemic disease. A protein that comigrates with the major form of pp41 (pp41A) and contains P-Tyr is also found in murine fibroblasts and B-lymphoid cells transformed by Abelson murine leukemia virus, which encodes the v-abl protein, and in platelet-derived growth factor-treated fibroblasts, in which it has been described previously. We analyzed three pairs of Epstein-Barr virus-immortalized B-cell lines from individual CML patients and found that only the lines in which active p210bcr-abl was present contained detectable pp41. We also performed immunoblotting with anti-P-Tyr antibodies on the same CML cell lines and detected at least four other putative substrates of p210bcr-abl, which were undetected with use of the two-dimensional gel technique.
Irradiated mice reconstituted with bone marrow cells infected with a retrovirus carrying the bcr-abl oncogene of human chronic myeloid leukemia are subject to a range of neoplastic hematopoietic diseases, both myeloid and lymphoid. Comparison of DBA/2 and C57BL/6 mice has revealed a marked strain difference in susceptibility to the various tumor types. The present study, performed with BALB/c mice, indicates that the kinetics and nature of the induced disease can be modulated by the infection procedure, as well as the genetic background, and that retroviral regulatory sequences may influence the outcome. A distinctive clonal myeloproliferative disorder, somewhat akin to chronic myeloid leukemia but with prominent erythroid and mast cell components, as well as granulocytic excess, was characterized.
Integrated Friend murine leukemia virus copies were analyzed by the Southern blotting procedure in myeloblastic cell lines obtained after in vitro infection of long-term mouse bone marrow cultures. Several steps leading to the generation of malignant myeloblastic cells after a long latency period were observed in the evolution of infected cultures. Shortly after infection, a random distribution of integrated provirus copies was observed in the DNA of normally differentiating myeloid cells. In contrast, a distinct pattern of integrated Friend murine leukemia virus copies was evident in the first non-differentiating immature myeloblastic cells appearing in cultures, suggesting a monoclonal origin of these cells. For each cell line, characteristic hybridizing fragments were conserved during the 1-year culture period necessary for the acquisition of tumorigenic properties and were also observed in tumors grafted in vivo. We can conclude that monoclonality is effective very early in the myeloid transformation process, as soon as the precursor cells are blocked in their differentiation.
The translocation of the c-abl oncogene from chromosome 9 to the bcr gene on chromosome 22 in cases of Philadelphia chromosome-positive chronic myelogenous leukemia (CML) generates an aberrant bcr-abl fusion transcript which may be intimately related to the pathogenesis of CML. Because factors controlling normal bcr expression might also be involved in the expression of this aberrant bcr-abl transcript, we studied the patterns of expression of the normal bcr gene in different cell types. We found that the normal bcr gene was expressed in many different types of human cells. Moreover, the bcr gene was evolutionarily conserved, and homologous bcr genomic sequences and RNA transcripts were readily detected in chick tissue. The highest level of bcr expression in chick tissue was in brain tissue, the lowest level was in liver tissue, and a truncated bcr mRNA was noted in chick testes. Normal bcr transcripts, in addition to the aberrant bcr-abl hybrid transcripts, have been found in all Philadelphia chromosome-positive CML cells studied to date. Within a given CML sample, the relative amounts of normal bcr RNA and aberrant bcr-abl RNA were similar. In addition, the normal bcr and the aberrant bcr-abl hybrid transcripts demonstrated similarly prolonged half-lives compared with that of the normal abl-related transcripts in CML cells. These findings suggest that in CML cells, similar cellular mechanisms control the steady-state levels of both the normal bcr and the bcr-abl fusion RNAs.
Two forms of activated BCR/ABL proteins, P210 and P185, that differ in BCR-derived sequences, are associated with Philadelphia chromosome-positive leukemias. One of these diseases is chronic myelogenous leukemia, an indolent disease arising in hematopoietic stem cells that is almost always associated with the P210 form of BCR/ABL. Acute lymphocytic leukemia, a more aggressive malignancy, can be associated with both forms of BCR/ABL. While it is virtually certain that BCR/ABL plays a central role in both of these diseases, the features that determine the association of a particular form with a given disease have not been elucidated. We have used the bone marrow reconstitution leukemogenesis model to test the hypothesis that BCR sequences influence the ability of activated ABL to transform different types of hematopoietic cells. Our studies reveal that both P185 and P210 induce a similar spectrum of hematological diseases, including granulocytic, myelomonocytic, and lymphocytic leukemias. Despite the similarity of the disease patterns, animals given P185-infected marrow developed a more aggressive disease after a shorter latent period than those given P210-infected marrow. These data demonstrate that the structure of the BCR/ABL oncoprotein does not affect the type of disease induced by each form of the oncogene but does control the potency of the oncogenic signal.
Chronic myeloid leukemia (CML) is a myeloproliferative disease that results from the BCR-ABL gene-induced transformation of a primitive hematopoietic cell. This disease has been extensively studied, and, as a result, a very effective therapy has been developed: the tyrosine kinase inhibitors. Although, there is a significant knowledge about the intrinsic biology of CML cells, alterations in their bone marrow microenvironment are not yet completely understood. In this concise review, we summarized recent findings on the composition and function of the bone marrow microenvironment in CML, and their importance in the progression of the disease and treatment resistance.
The discovery of the breakpoint cluster (bcl)-Abelson kinase (abl) oncogene was one of the most important milestones for elucidating the pathophysiology of chronic myeloid leukemia (CML). Numerous studies have shown that due to various mutations, it has been difficult to find drugs that maintain significant efficacy over an extended period of time. The resistance of the BCR-ABL protein to pharmacotherapy is primarily due to point mutations and gene amplification, as well as a number of BCR-ABL—independent mechanisms. Tyrosine kinase inhibitors (TKIs), such as imatinib, have been shown to have significant efficacy in certain CML patients. However, the development of a mutation in the gate keeper region of BCR-ABL protein, “T315I,” significantly decreased or abrogated the clinical efficacy of certain TKIs. In this chapter, we discuss (1) the most common resistance mechanisms that occur in the BCR-ABL protein in CML patients and (2) the approaches that have been developed over the last two decades to inhibit the function of the BCR-ABL protein.
Classifying the hematological malignancies by assigning cells to their normal counterpart and describing the nature of disease progression are entirely reliant on an accurate picture for the development of the multifarious types of blood and immune cells. In recent years, our understanding of the complex relationships between the various hematopoietic stem cell-derived cell lineages has undergone substantial revision. There has been similar progress in how we describe the nature of the “target” cells that genetic insults transform to give rise to the hematological malignancies. Here I describe how both longstanding and new information has influenced classifying, for diagnosis, the hematological malignancies.
Chronic myeloid leukemia (CML) is a hematopoietic stem cell disorder caused by constitutively active BCR-ABL1 tyrosine kinase resulting from the t(9;22) Philadelphia translocation. Imatinib, a BCR-ABL1 tyrosine kinase inhibitor (TKI), is a revolutionary molecular target inhibitor for CML. However, leukemic stem cells (LSCs) eventually become resistant to imatinib and thereby cause relapse. The next generation BCR-ABL1 TKI dasatinib is also unable to eliminate CML LSCs. On the other hand, the third generation BCR-ABL1 TKI ponatinib is not well-studied in terms of its efficacy on CML LSCs. Here, we evaluate the efficacy of ponatinib against CML LSC-containing lin-Sca-1+c-Kit+ (LSK) cells using a mouse CML-like model. To this end, we compared the efficacy of imatinib, dasatinib and ponatinib on CML LSK cells and showed that ponatinib is more effective at eliminating CML LSK cells. Our results suggest that ponatinib could be potentially useful for achieving treatment-free remission in CML patients.
The heteromorphic regions of chromosome 22 were studied with the quinacrine mustard fluorescence technique in eight patients with Philadelphia chromosome-positive chronic myelocytic leukemia and in their parents. The fluorescence pattern showed that the Philadelphia chromosome had originated from the paternal chromosome 22 in one case and the maternal in another. The other six cases were noninformative in this respect. The fluorescence pattern was consistent between cells in both the informative cases. These results speak in favor of a clonal origin of the Philadelphia chromosome from either the paternal or the maternal chromosome 22.
Bone marrow aspirates from 12 patients with malignant hematologic disorders and abnormal bone marrow karyotypes were used to establish in vitro fibroblast cultures. The karyotypes of such fibroblasts were consistently normal. This finding is at variance with results presented by others. The origin of bone marrow derived fibroblasts remains obscure.
Terminal deoxynucleotidyl transferase, an enzyme which catalyzes the polymerization of deoxyribonucleoside triphosphates, elongating oligo- or polydeoxynucleotide chains, but without direction from a nucleic acid template, is thought to be specific for thymus gland and thymus- derived cells. We have confirmed the observations that high levels are characteristic of thymus gland with both human and calf tissue and that elevated levels may be found in some cases of acute lymphocytic leukemia. High levels were also found in human lymphoblast cell lines with T-cell characteristics, and insignificant activity was observed in leukocytes of patients with chronic myelogenous leukemia not in acute blast phase of the disease, chronic lymphocytic leukemia, human B- cells, and normal human blood lymphocytes even after stimulation with phytohemagglutinin. However, high levels (approximately 200 nmoles/hr/10(9) cells) equivalent to those in thymus tissue and lymphoblast cell lines with T-cell characteristics were found in the peripheral blood blast cells of four patients with chronic myelogenous leukemia in an acute blast phase of their disease. One hypothesis that may explain the present results is that in chronic myelogenous leukemia in acute blast phase of the disease the proliferative blast response may not always be myeloblasts but in some cases it may be lymphoblasts.
An attempt was made to determine the origin of bone marrow fibroblasts which almost always appear when bone marrow aspirates and explants are grown on solid surfaces in tissue culture. Bone marrow aspirates from an individual with chronic myelogenous leukemia demonstrating the Ph-1 chromosome and from an individual with acute leukemia demonstrating C-group trisomy in marrow cells served as sources of the fibroblasts. The fibroblasts that grew from these aspirates were analyzed for the presence of these chromosome markers found in the leukemic cells. The positive findings of the Ph-1 chromosome and C-group trisomy in a proportion of the fibroblasts indicates that at least some leukemic bone marrow fibroblasts are derived from hemopoietic cells. The origin of the bone marrow fibroblasts are derived from hemopoietic somes remains unknown. They could be derived from (1) hemopoietic cells cytogenetically unaffected by the leukemic process, (2) stromal elements of the bone marrow, or (3) both cytogenetically unaffected hemopoietic cells and stromal elements.
Thirteen patients with chronic myelogenous leukemia continued to have the Ph1 chromosomes in 90-100 per cent dividing marrow cells during drug-induced clinical remissions. The Ph1 chromosome was present in erythroid as well as granulocytic marrow cells, and possibly in megakaryocytes.
The presence of Ph1 chromosomes was also studied in cultures of peripheral blood. In six patients in relapse, 40 per cent of metaphases contained the Ph1 chromosome, and the percentage of these cells corresponded roughly to the relative frequency of immature granulocytes in the blood. In contrast, during remission, few or no Ph1 chromosomes were found in peripheral blood cultures, presumably because in the absence of immature granulocytes the dividing cells in the cultures originate from lymphocytes, as they do in normal blood.
It is suggested that the Ph1 chromosome usually arises in a precursor cell common to the erythroid, granulocytic, and megakaryocytic, but not the lymphoid series of hemopoietic cells.
A male patient with CML had an unusual No. 9 chromosome with a typical Ph¹ chromosome translocation. This unusual No. 9 chromosome showed heteromorphism at the 9qh region and a missing band in the middle of its long arm. These characteristic features of the leukemic cells strongly support the monoclonal origin of CML.
A male patient aged 69 yr with chronic myeloid leukaemia was a constitutional XY/XXY sex chromosomal mosaic as indicated by a positive sex chromatin in neutrophils and cytogenetic studies of lymphocytes cultured with phytohaemagglutinin. He was of normal phenotype and intelligence. In a bone-marrow aspirate taken when the patient was in the acute phase of leukaemia the 46, XY cell line carried the Ph1 chromosome, whereas the 47, XXY cell line did not. Two further cell lines were considered to have been derived by clonal evolution of the 46,XY,Ph1-positive line, although one of them possessed ambiguous features. The results support a clonal origin of the Ph1 chromosome, and presumably also of chronic myeloid leukaemia. An interesting feature of the bone marrow was that the 46, XY, Ph1-positive cell line had apparently replaced the normal 46, XY cells but not the 47, XXY cells.
In a female patient with chronic myelocytic leukemia (CML) the translocation 9q +; 22q was constantly found to involve only the variant chromosome 9 with an unusually long secondary constriction. The finding indicates a unicellular origin of CML.
Two women with polycythemia vera and heterozygosity (GdB/GdA) at the X-chromosome-linked locus for glucose-6-phosphate dehydrogenase were studied to determine the nature of the cellular origin of their polycythemia. In contrast to unaffected tissue, such as skin fibroblasts, which consisted of both B and A types, the glucose-6-phosphate dehydrogenase of the patients' erythrocytes, granulocytes and platelets was only of Type A. These results provide direct evidence for the stem-cell nature of polycythemia vera and strongly imply a clonal origin for this disease. The fact that no descendants of the presumed normal stem cells were found in circulation suggests that bone-marrow proliferation in this disorder is influenced by local (intramarrow) regulatory factors.
Peripheral leukocytes from patients with and without leukemia were assayed for presence of terminal deoxynucleotidyl transferase. Activity of this enzyme was detected in circulating leukemic cells from 11 to 13 patients with acute lymphoblastic leukemia, and in one of four with chronic myelogenous leukemia in blast crisis, but not in leukocytes from patients with other kinds of leukemia or in normal leukocytes. Its presence in a patient with chronic myelogenous leukemia in blast crisis lends biochemical support to the suggestion that some patients with chronic myelogenous leukemia undergo a lymphoblastic rather than a myeloblastic crisis. The thymocyte and leukemic-cell enzyme have the same substrate and primer preference. Normal thymocytes and leukemic cells contain two forms of terminal deoxynucleotidyl transferase that can be separated by phosphocellulose chromatography. The enzyme may provide a means for classifying leukemic cells on a biochemical basis independently of classic morphologic and clinical criteria.
Terminal deoxynucleotidyl transferase, an enzyme which catalyzes the polymerization of deoxyribonucleoside triphosphates, elongating oligo- or polydeoxynucleotide chains, but without direction from a nucleic acid template, is thought to be specific for thymus gland and thymus-derived cells. We have confirmed the observations that high levels are characteristic of thymus gland with both human and calf tissue and that elevated levels may be found in some cases of acute lymphocytic leukemia. High levels were also found in human lymphoblast cell lines with T-cell characteristics, and insignificant activity was observed in leukocytes of patients with chronic myelogenous leukemia not in acute blast phase of the disease, chronic lymphocytic leukemia, human B-cells, and normal human blood lymphocytes even after stimulation with phytohemagglutinin. However, high levels (approximately 200 nmoles/hr/10(9) cells) equivalent to those in thymus tissue and lymphoblast cell lines with T-cell characteristics were found in the peripheral blood blast cells of four patients with chronic myelogenous leukemia in an acute blast phase of their disease. One hypothesis that may explain the present results is that in chronic myelogenous leukemia in acute blast phase of the disease the proliferative blast response may not always be myeloblasts but in some cases it may be lymphoblasts.
CELLS from nine consecutive patients with chronic myelogenous leukaemia (CML) have been analysed with quinacrine fluorescence and various Giemsa staining techniques. The Philadelphia (Ph1) chromosome in all nine patients represents a deletion of the long arm of chromosome 22 (22q-)1,2. An unsuspected abnormality in all cells from the nine patients has been detected with these new staining techniques. It consists of the addition of dully fluorescing material to the end of the long arm of one chromosome 9 (9q+). In Giemsa-stained preparations, this material appears as an additional faint terminal band in one chromosome 9. The amount of additional material is approximately equal to the amount missing from the Ph1 (22q-) chromosome, suggesting that there may be a hitherto undetected translocation between the long arm of 22 and the long arm of 9, producing the 9q+ chromosome.
Following busulphan‐induced bone marrow insufficiency a patient with chronic granulocytic leukaemia has had a remission lasting 10 yr. During this time repeated chromosome studies on bone marrow have shown the persistence of a majority of Ph1 negative cells. The relationship of this finding to the unusually long remission is discussed.
This article has no abstract; the first 100 words appear below.
BONE-marrow cells from the majority of patients with chronic myelocytic leukemia have a specific and characteristic aberration of chromosome 22, called the Philadelphia (Ph¹) chromosome.¹ This abnormality, which may be found even before the onset of overt leukemia,² is present throughout the course of clinical disease. Although the Ph¹ chromosome probably is important in the pathogenesis of the disease the primary causes (which presumably are also the factors inducing the chromosome abnormality) remain largely unknown. An indirect approach to elucidating causative factors involves determining the number of cells from which chronic myelocytic leukemia arises. Single-cell (clonal) origin would be . . .
Supported by a grant (GM 15253) from the National Institutes of Health, U.S. Public Health Service.
We are indebted to Professor W.F.M. Fulton for allowing us to study this patient and to Mr. G. Herner for technical assistance.
Source Information
From the Haematology Unit, Department of Pathology, University of Aberdeen, Scotland, the Department of Medicine, University of Nairobi, Kenya, and the departments of Medicine and Genetics, University of Washington, Seattle, Wash. (address reprint requests to Dr. Barr at the Haematology Unit, Department of Pathology, University of Aberdeen, University Medical Bldgs., Foresterhill, Aberdeen AB9 2ZD, Scotland).
Cytogenetic studies on a 7�--yr-oId child pre- senting with clinical and hematological fea- tures of chronic myeloid leukemia (CML) re- vealed a constitutional 46 XV/47 XVV mosaicism in skin, blood, and marrow. A third 46 XV, Ph1 cell line predominated in the marrow on initial presentation and in subsequent acute transformation. Assessment of granulocytic colony-forming capacity in agar culture revealed that colony-forming cells (CFCs) were greatly increased in the circulation and possessed the abnormal light buoyant density and low susceptibility to tritiated thy- midine killing which distinguishes leukemic CFCs from normal. During acute transforma- tion colony-forming capacity was lost but small, poorly differentiated cell clusters persisted in culture. Only 46 XV, Ph' metaphase were ob- tained following cytogenetic analysis of in vitro colonies and clusters displaying leukemic growth characteristics in agar culture. The co- existence of CML and sex chromosome mosa- icism in this patient provides further support for the uniclonal origin of CML and indicates that cytogenetic instability implicated in mosaicism may carry an increased risk of further cyto- genetic evolution with emergence of the Ph' chromosome.
Karyotypic analysis was performed on agar cultures of blood or bone marrow from 12 patients with acute or chronic myeloid or myelomonocytic leukemia in whom karyotypic markers were present. The granulocytic colonies and clusters which developed on culture were shown to be derived from representative cells of the leukemic population. In two patients with acute leukemia in remission, normal colonies with a normal karyotype were grown from marrow cells but in two patients with chronic myeloid leukemia in remission the Ph1 abnormality persisted in colony cells. The agar culture technique appears to be ideal for following the emergence and disappearance of leukemic and normal granulopoietic populations in patients with these types of leukemia.
Current evidence suggests that chronic myeloid leukemia (CML) must be considered a clonal disease of a pluripotent hematopoietic stem cell compartment. The Philadelphia chromosome (Ph1) abnormality (translocation of a portion of the long arms of chromosome 22 onto chromosome 9) is found not only in neutrophil precursors, but in precursors of red cells, eosinophils, probably platelets and monocytes. These observations plus the failure to find the Ph1 in skin cells or lymphocytes from patients with CML or in bone marrow cells from their nonleukemic identical twins indicate involvement of selected cellular systems. Blastic crisis, the most common cause of death in patients with CML, is characterized by increasing failure of maturation with continued overproduction of immature cells, most commonly myeloblasts and promyelocytes, but underproduction of mature neutrophils, platelets and red cells. Based on chromosome studies, blast crisis appears to be due to acquisition of yet another clone of abnormal stem cells in addition to the Ph1 clone. Patients going into blast crisis have been observed to acquire a second Ph1 or other abnormal chromosome, lose the second with a return to a CML like phase following therapy, and reacquire it as blastic crisis recurs. The morphologic expression is quite variable, and the predominant cell may be a myeloblast, promyelocyte, eosinophilic promyelocyte, basophilic promyelocyte, monocyte, 'myelomonocyte' or proerythroblast. The suggestion is raised that still another form of blastic conversion may occur, one that does not seem to make sense at first glance, namely, lymphoblastic conversion in CML.
The relative compositions of A and B types of G 6 PD have been estimated in erythrocytes, granulocytes, lymphocytes, skin and skeletal muscle, obtained from 42 Gd(B)/Gd(A) heterozygotes. The high positive correlations found between the various studied tissues are taken to strongly suggest a common precursor pool, the size of which is estimated to be 16 (10-25) cells. This number could represent progenitor cells for mesoderm or a more primordial cell pool such as one for ectoderm (from which mesoderm subsequently differentiates) or even the total number of embryoblasts present at the time of X chromosome inactivation. Thus, a minimum estimate for the number of embryoblasts present in man at the time of X inactivation is 16 (10-25). The data also suggest that when tissue specific clonal development occurs, the number of cells entering each pathway is relatively large ( > 80 cells) and that the subsequent growth of skin and skeletal muscle is characterized by considerable cell mixing.
Involvement of the erythroid series in chronic myeloid leukaemia may be evidenced haematologically by a normochromic anaemia with anisocytosis, the presence of circulating normoblasts, the presence of bizarre normoblasts in the marrow and peripheral blood, and rarely the presence of sideroblasts. The Philadelphia (Ph1) chromosome has been assumed to be present in the erythrocyte as well as the granulocyte series on good but circumstantial evidence and it is generally agreed that the leukaemic process probably affects the basic stem cell of the red cells, granulocytes and platelets.
When blastic crisis occurs there is an increase in the percentage of blast cells in marrow and peripheral blood, and anaemia, thrombocytopenia and resistance to therapy accompany the apparently irreversible downward clinical course. Aneuploid cell lines emerge (Hammouda, Quaglino and Hayhoe, 1964). We are here presenting evidence that aneuploid Ph1-positive cell lines are also found in the erythroid cells in blastic crisis and therefore that when this occurs the essential change may also affect the basic stem cell. In a case of chronic myeloid leukaemia seen recently, a sideroblastic phase was found terminally and it was possible to combine standard cytogenetic methods with Perls's prussian blue stain for iron and demonstrate siderotic granules in the cytoplasm surrounding Ph1-positive aneuploid chromosome spreads. We thus prove the erythroid nature of these spreads, and incidentally provide more direct evidence than offered hitherto, for the presence of the Ph1 chromosome in the erythroid cells.
In a study of 41 patients with chronic myelocytic leukemia, two were found to have the 6-phosphogluconate dehydrogenase heterozygous phenotype A-B, and two had the phenotype characteristic of Pd(B) homozygosity. Since one of the two with Pd(B) homozygosity was the mother of two children with the A phenotype, it was presumed that she carried a Pd(A) gene not expressed in her blood cells. his was confirmed by electrophoretic analysis of her fibroblasts, which had the A-B phenotypic pattern. Gene deletion is considered to be the most likely explanation.
GRANULOCYTES, erythroblasts and megakaryocytes from patients with chronic granulocytic leukaemia contain the Ph1 chromosome1-3 in contrast to lymphocytes, which do not3. Skin fibroblasts from at least one case of chronic granulocytic leukaemia did not, however, have the Ph1 chromosome4. Because myelofibrosis occurs in association with chronic granulocytic leukaemia5-7 and because it is also considered to be a myeloproliferative disorder, the question has arisen7 of the presence of a Ph1 chromosome in marrow fibroblasts.
An attempt was mad to determine the origin of bone marrow fibroblasts which almost always appear when bone marrow aspirates and explants are grown on solid surfaces in tissue culture. Bone marrow aspirates from two individuals with chronic myelogenous leukemia demonstrating the Ph-1 chromosome and from an individual with acute leukemia demonstrating the trisomic C-group karyotype served as sources of the fibroblasts. These were analyzed for the presence of the marker chromosomes found in the leukemic cells of these patients. Over half of the dividing fibroblasts demonstrated the marker chromosomes. This positive finding indicates that at least some bone marrow fibroblasts are derived from hemopoietic stem cells. These cells should, therefore, prove useful in the study of cellular differentiation.
Human leukemic cells with a marker (Philadelphia; Ph(1)) chromosome gave rise to granulocytic and mononuclear cell colonies when grown in vitro. All metaphases from a single colony were either Ph(1) positive or Ph(1) negative. No colonies contained a mixed cell population. This suggests that leukemic and normal cells exist simultaneously and that in vitro colonies are clonal in origin.
Bone marrow (about 0.5 ml) from au erythropoietic region is freed of blood clots by washing 1-3 min in 1 μg/ml colchicine solution (2-3 ml) and then soaking 1-2 hr at 20-30° C in a second change. For mammalian or avian marrows, the colchicine is made up in phosphate-buffered (pH 7) physiological NaCl solution; for amphibian, Ringer's A solution. Next the specimens are soaked about 20 min in a hypotonic solution as follows: for mammalian, 1% Na-citrate; for avian, a 1:4 dilution of the buffered NaCl solution by distilled water; and for amphibian, Ringer's A-distilled water, 1:1. Then they are heated in a mixture of 2% orcein in 45% acetic acid and 1 N HCl, 9:1. Immediately after heating, squash preparations are made with 2% acetic-orcein in the usual manner. An alternative method is to dissociate the marrow cells by agitating after colchicine treatment. Then, recovering the cells between changes by low-speed centrifugation, to carry out the hypotonic treatment and subsequent fixation in Carnoy's solution I (alcohol acetic, 3:1) before drying the cells onto slides from the fixative. After thorough drying the slides may be stained 10-20 min in acetic orcein, or by other suitable technics.
The frequency of the Ph1 chromosome in freshly aspirated marrow cells of 14 patients with typical chronic myelocytic leukemia processed by a "direct technic" without resort to culture or colchicine was significantly higher (> 75 per cent) than that observed in the cultured blood cells (< 35 per cent) of the same subjects. The karyotypic abnormally of the abbreviated G-group chromosome would appear not to be related to therapy, since the frequency with which it occurred was not materially affected by treatment (including radiation). The Ph1 chromosome was not observed in any of the metaphases of blood or marrow of 12 subjects who had developed a leukemia-like picture complicating either myelofibrosis, polycythemia vera or myeloid metaplasia. A new chromosome abnormality—a shortened D-group chromosome—was observed with about the same frequency in the blood and marrow metaphases of a female patient with treated chronic myelocytic leukemia. This new karyotypic abnormality was associated with the highest frequency of the Ph1 chromosome in cultured blood cells in the group studied. The Ph1 chromosome was observed in the metaphases of a patient with the blastic phase of chronic myelocytic leukemia. The variations of the morphology of the Ph1 chromosome are discussed and illustrated, especially in relation to the Y-chromosome. In four patients with an atypical picture of CML, the Ph1 chromosome was not observed either in the marrow or cultured blood.
Idiopathic myelofibrosis: stem cell abnormality and probable neoplastic origin
- Jacobson
Jacobson RJ, Fialkow PJ: Idiopathic myelofibrosis: stem cell abnormality and probable neoplastic origin. Clin Res 24: 439A, 1976.
6-Phosphogluconate dehydrcgenase: hemizygous manifestation in a patient with leukemia Genetic markers in chronic myelocytic leukemia: evidence opposing autosomal 18
- Fialkow Pj
- R Lisker
- J Detter
Fialkow PJ, Lisker R, Detter J, et al.: 6-Phosphogluconate dehydrcgenase: hemizygous manifestation in a patient with leukemia. Science 163: 194, 1969. Fialkow PJ, Lisker R, Giblett ER, et al.: Genetic markers in chronic myelocytic leukemia: evidence opposing autosomal 18. CHRONIC MYELOCYTIC LEUKEMIA-FIALKOW ET AL 19. 20. 21. 22. 23. 24. 25. 26. 27.