Malika Kapadia

Casein Kinase II (CK2), Glycogen Synthase Kinase-3 (GSK-3) and Ikaros mediated regulation of leukemia

Signaling networks that regulate cellular proliferation often involve complex interactions between several signaling pathways. In this manuscript we review the crosstalk between the Casein Kinase II (CK2) and Glycogen Synthase Kinase-3 (GSK-3) pathways that plays a critical role in the regulation of cellular proliferation in leukemia. Both CK2 and GSK-3 are potential targets for anti-leukemia treatment. Previously published data suggest that CK2 and GSK-3 act synergistically to promote the phosphatidylinositol-3 kinase (PI3K) pathway via phosphorylation of PTEN. More recent data demonstrate another mechanism through which CK2 promotes the PI3K pathway - via transcriptional regulation of PI3K pathway genes by the newly-discovered CK2-Ikaros axis. Together, these data suggest that the CK2 and GSK-3 pathways regulate AKT/PI3K signaling in leukemia via two complementary mechanisms: a) direct phosphorylation of PTEN and b) transcriptional regulation of PI3K-promoting genes. Functional interactions between CK2, Ikaros and GSK3 define a novel signaling network that regulates proliferation of leukemia cells. This regulatory network involves both direct posttranslational modifications (by CK and GSK-3) and transcriptional regulation (via CK2-mediated phosphorylation of Ikaros). This information provides a basis for the development of targeted therapy for leukemia.

Protein signaling and regulation of gene transcription in leukemia: role of the Casein Kinase II-Ikaros axis

Protein signaling and regulation of gene expression are the two major mechanisms that regulate cellular proliferation in leukemia. Discerning the function of these processes is essential for understanding the pathogenesis of leukemia and for developing the targeted therapies. Here, we provide an overview of one of the mechanisms that regulates gene transcription in leukemia. This mechanism involves the direct interaction between Casein Kinase II (CK2) and the Ikaros transcription factor. Ikaros (IKZF1) functions as a master regulator of hematopoiesis and a tumor suppressor in acute lymphoblastic leukemia (ALL). Impaired Ikaros function results in the development of high-risk leukemia. Ikaros binds to the upstream regulatory elements of its target genes and regulates their transcription via chromatin remodeling. In vivo, Ikaros is a target for CK2, a pro-oncogenic kinase. CK2 directly phosphorylates Ikaros at multiple amino acids. Functional experiments showed that CK2-mediated phosphorylation of Ikaros, regulates Ikaros’ DNA binding affinity, subcellular localization and protein stability. Recent studies revealed that phosphorylation of Ikaros by CK2 regulates Ikaros binding and repression of the terminal deoxytransferase (TdT) gene in normal thymocytes and in T-cell ALL. Available data suggest that the oncogenic activity of CK2 in leukemia involves functional inactivation of Ikaros and provide a rationale for CK2 inhibitors as a potential treatment for ALL.

Pulmonary Complications Associated with HSCT

While outcomes for patients who undergo allogeneic hematopoietic stem cell transplantation (HSCT) have improved over the past 10–20 years, pulmonary complications after allogeneic HSCT remain a leading cause of morbidity and mortality. Overall, 25–50% of pediatric HSCT patients will develop pulmonary complications. Thus, prevention, early detection, and intervention are key to minimizing the sequelae from HSCT-associated pulmonary complications. HSCT-associated pulmonary complications can be classified as infectious or noninfectious, and they often follow a predictable timeline, occurring during discrete phases of HSCT (pre-engraftment, early post-engraftment, late post-engraftment). However, certain post-HSCT pulmonary complications span the entire post-HSCT course. The most common causes of noninfectious pulmonary complications are related to the conditioning regimen used which can result in varying degrees of acute or delayed lung injury, the degree of recipient–donor HLA histoincompatibility, the hematopoietic stem cell (HSC) source, the degree of graft manipulation, and the development of graft-versus-host disease (GvHD), both acute and chronic. Infectious etiologies can be caused by any class of pathogen including bacterial, viral, fungal, and protozoan. They usually occur during periods of profound and/or prolonged neutropenia and/or impaired or delayed cellular and humoral immune recovery. Immunosuppression used to prevent or treat GvHD also places a HSCT recipient at high risk for developing pulmonary infections that can be life-threatening. This chapter discusses the most common pulmonary complications associated with HSCT by time period post-HSCT.

How to Select a Donor and Hematopoietic Stem Cell Source: Related Versus Unrelated Donors for Allogeneic HSCT

The selection of the most suitable donor and stem cell source is a critical component of hematopoietic stem cell transplantation (HSCT). The factors that contribute to this selection are many, making the process complex. The most important contributing factor of donor selection and stem source is based on the inherent genetic makeup of the donor as it relates to the HSCT recipient. The cluster of genes that compared for compatibility (termed histocompatibility) contain human leukocyte antigens (HLA) genes located in the short arm of chromosome 6 in humans. These genes are inherited together (i.e., linked), and the most important determinants are HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DQB1, and HLA-DPB1. Because a person inherits one HLA cluster (termed locus) from each parent, siblings with the same parents have a 25% chance to be a “match” with the potential HSCT recipient. The process of evaluating histocompatibility is termed HLA matching and can be performed at the antigen and allele level. The allele level is more accurate and thus is termed “high-resolution” typing. Because increased HLA disparity between the donor and recipient increases post-HSCT morbidity and mortality, every effort is made to select the most suitable donor in terms of HLA histocompatibility. Potential hematopoietic stem cell (HSC) sources include bone marrow, peripheral blood after cytokine mobilization, and umbilical cord blood. Each HSC source is commonly used, with bone marrow as the most common source for pediatric allogeneic HSCT. However, umbilical cord blood is used frequently as an alternative source when no suitable related or unrelated matched sibling donor is available. Because of the nature of the naive T cells contained within umbilical cord blood, greater HLA disparity between umbilical cord blood donor source and the potential HSCT recipient is better tolerated than with bone marrow or peripheral blood HSCs. Haploidentical related donors as HSC source is being used more frequently but is still considered investigational in the pediatric population.

Renal Toxicities in the Peri-HSCT Period

Renal dysfunction within the first 100 days post-hematopoietic stem cell transplantation (HSCT) is common. Many HSCT patients have some degree of renal dysfunction prior to HSCT, and the degree of this dysfunction impacts the extent of renal dysfunction that can occur during the pre-engraftment and early post-engraftment periods of the HSCT course. Renal dysfunction may be due to renal tubule damage, compromised renal blood flow, and obstruction or irritation of post-renal structures. Renal dysfunction impairs the ability of the kidneys to maintain acid-base and electrolyte balance as well as maintain fluid balance and eliminate waste. The renal complications post-HSCT are discussed in Chap. 22. The most common renal toxicities are related to medications frequently used during the peri-HSCT period. These include calcineurin inhibitors, antifungal agents, antibiotics (particularly aminoglycosides), and antiviral agents. In addition, components of conditioning regimens such as alkylators and irradiation to the bladder can cause significant renal toxicity. An infrequent but very serious renal complication is hemorrhagic cystitis which is the focus of this chapter. The remainder of HSCT-associated renal complications is addressed in Chap. 22.

Intracranial malignant peripheral nerve sheath tumor variant: an unusual neurovascular phenotype sarcoma case invading through the petrous bone

IntroductionIntracranial malignant peripheral nerve sheath tumor (MPNST) is exceedingly rare. Previously reported cases of intracranial MPNST have been associated with development within a prominent cranial nerve.MethodsThis is the first report of an MPNST with both nerve sheath and vascular phenotype that follows the neurovascular bundle, without arising in a major cranial nerve or in the setting of neurofibromatosis type 1 (NF1).ResultsThe patient is a 14-year-old boy with a history of worsening headaches for the past several months, left-sided hearing loss, nausea, vomiting, and vertigo. MRI was performed that demonstrated a large extra-axial tumor compressing the left infratemporal posterior temporal region. The tumor was associated with significant destruction of the superior portion of the petrous bone and extension through the petrous into the upper posterior fossa, immediately below the tentorium. The patient underwent surgical debulking and adjuvant chemotherapy with doxorubicin and ifosfamide. Pathology demonstrated a variant malignant peripheral nerve sheath tumor with both nerve sheath and vascular phenotype by immunostains. The patient’s symptoms improved following treatment.ConclusionWe present the first reported case of an intracranial MPNST variant that developed along the neurovascular bundle as a sarcoma with both nerve sheath and vascular phenotype through the petrous bone and not associated with a major cranial nerve or with stigmata of neurofibromatosis type 1 (NF1). Although this is an extremely unusual presentation due to location and lack of prominent cranial nerves in that location, it is not unusual for benign nerve sheath tumors to follow the neurovascular bundle through foramen of cortical long bone or pelvis. This case suggests that physicians should incorporate intracranial MPNST variant into their differential diagnosis in the cranium, even when tumor is not located near a prominent cranial nerve. Surgical debulking and adjuvant chemotherapy with doxorubicin and ifosfamide has led to improvement in patient symptoms.

Abstract 5507:RAG1high expression associated withIKZF1dysfunction in adult B-cell acute lymphoblastic leukemia

The recombination activating gene (RAG)-mediated recombination is the dominant mutational process and the predominant driver of oncogenic genomic rearrangement in acute lymphoblastic leukemia (ALL). This then leads to further leukemic clonal evolution. IKZF1 encodes a kruppel-like zinc finger protein, IKAROS that is essential for normal hematopoiesis and acts as a tumor suppressor in ALL. The genetic defects of a single IKZF1 allele are linked to the development of human ALL, characterized by an increased risk of relapses and poorer prognosis. We observed that RAG1 is significantly increased in subsets of B-ALL patients. High RAG1 expression correlates with high proliferation markers. IKAROS directly binds to the RAG1 promoter in B-ALL cells by quantitative chromatin-immunoprecipitation assay. IKAROS suppresses RAG1 promoter activity by luciferase reporter assay. Lentiviral IKAROS expression significantly suppresses RAG1 expression, but IKAROS shRNA promotes RAG1 expression in B-ALL cells. CK2 inhibitor by restoring IKAROS activity, significantly suppresses RAG1 expression in an IKAROS-dependent manner in B-ALL cells. RAG1 expression is significantly higher in patients with IKZF1 deletion, as compared to patients without IKZF1 deletion. Treatment with CK2 inhibitor also results in an increase in IKZF1 binding to the RAG1 promoter and suppression of RAG1 expression in primary B-ALL cells. Taken together, our results demonstrate that high expression of RAG1 correlates with high proliferation markers in B-ALL, and are the first to demonstrate that IKAROS directly suppresses RAG1 expression. Our data suggest RAG1 high expression works together with IKAROS dysfunction to drive oncogenesis of B-ALL, which have significance in an integrated prognostic model for adult ALL. Citation Format: zheng Ge, Qi Han, Jinlong Ma, Yan Gu, Huihui Song, Malika Kapadia, Sinisa Dovat, Chunhua Song. RAG1 high expression associated with IKZF1 dysfunction in adult B-cell acute lymphoblastic leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5507.

Abstract 5542: Regulation of cell cycle control in T-cell acute lymphoblastic leukemia by Ikaros and Casein Kinase II

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy that represents a therapeutic challenge. Next-generation sequencing revealed that a subset of T-ALL harbors inactivating mutations or deletion of one allele of the IKZF1 tumor suppressor. These data suggest that IKZF1 acts as a tumor suppressor in T-ALL. The IKZF1 gene encodes the Ikaros protein that functions as a regulator of transcription and a tumor suppressor in B cell acute lymphoblastic leukemia. However, the molecular mechanism of Ikaros tumor suppressor function in T-ALL is unclear. Using quantitative chromatin immunoprecipitation (qChIP), we determined that Ikaros binds to the promoter regions of the CDC2 and CDC7 cell cycle genes in primary T-ALL cells in vivo. Gain-of function experiments showed that Ikaros overexpression in T-ALL results in reduced expression of CDC2 and CDC7, as evidenced by quantitative RT-PCR (qRT-PCR) and Western blot. The knock-down of Ikaros with shRNA in T-ALL cells resulted in increased transcription of CDC2 and CDC7 as indicated by qRT-PCR. These data suggest that Ikaros can regulate cell cycle progression in T-ALL by repressing transcription of the CDC2 and CDC7 genes. Next, we studied the mechanisms that regulate Ikaros’ ability to repress CDC2 and CDC7 in T-ALL. Ikaros function as a transcriptional repressor is regulated by Casein Kinase II (CK2). CK2 is overexpressed in hematopoietic malignancies and increased expression of CK2 results in T-ALL in murine models. We tested the effect of CK2 inhibition on Ikaros’ ability to regulate transcription of CDC2 and CDC7 in human T-ALL. Molecular inhibition of CK2 with shRNA against the CK2 catalytic subunit resulted in reduced transcription of CDC2 and CDC7, as evidenced by qRT-PCR. This was associated with increased DNA-binding of Ikaros to promoters of CDC2 and CDC7, as shown by qChIP. These data suggest that CK2 impairs Ikaros’ ability to transcriptionally repress CDC2 and CDC7 and to regulate cell cycle progression in T-ALL. Inhibition of CK2 enhances transcriptional repression of CDC2 and CDC7 by Ikaros, resulting in improved control of cell cycle progression in T-ALL. In conclusion, our results show that control of cell cycle progression in T-ALL occurs trough Ikaros-mediated transcriptional regulation of CDC2 and CDC7. Overexpession of CK2 impairs Ikaros ability to repress CDC2 and CDC7 expression, which contributes to deregulation of cell cycle control in T-ALL. Results suggest a potential mechanism of therapeutic action of CK2 inhibitors for the treatment of T-ALL. Note: This abstract was not presented at the meeting. Citation Format: Mario A. Soliman, Tommy Hu, Malika Kapadia, Elanora Dovat, Yali Ding, Chunhua Song, Jonathon L. Payne, Sinisa Dovat. Regulation of cell cycle control in T-cell acute lymphoblastic leukemia by Ikaros and Casein Kinase II [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5542. doi:10.1158/1538-7445.AM2017-5542