What is Acute Lymphoblastic Leukemia?

Acute lymphoblastic leukemia (ALL) occurs as a result of abnormal accumulation of the lymphoid progenitor cells in the bone marrow, blood and extramedullary regions. Acute lymphoblastic leukemia consists of a B-cell precursor lineage (B-ALL) and a T-cell precursor lineage (T-ALL) subtypes. Acute lymphoblastic leukemia is subdivided into these subtypes based on morphological, immunophenotypic, cytogenetic and chromosomal properties. Both types are caused by structural chromosomal changes, changes in the number of copies in DNA, and sequence mutations that cause leukomogenesis. 80% of Acute lymphoblastic leukemia cases are seen in children, but the results are more severe when seen in adults.

Acute Lymphoblastic Leukemia

Acute lymphoblastic leukemia originates from malignant hematopoietic B- and T-lineage lymphoids that have genetic abnormalities including mutations, aneuploidies and translocations in the genes regulating cell growth, division, differentiation and other important cellular processes. Acute lymphoblastic leukemia is characterized by the accumulation of these malignant, immature lymphoid cells within the bone marrow, peripheral blood and extramedullary sites such as spleen. Common symptoms seen in Acute lymphoblastic leukemia include bone marrow related anemia, leukopenia, thrombocytopenia, fever, weight loss, easy bleeding, fatigue and brusing.

Diagnosis is carried out by detecting 20% or more lymphoblasts in the peripheral blood or bone marrow. To be able to confirm the diagnosis, morphological analysis, immunophenotyping, flow cytometry and cytogenetic analysis are commonly used. Although Acute lymphoblastic leukemia is the most commonly seen childhood leukemia (80%), it can be seen in adolescents and young adults with severe consequences and lower survival rates due to the heterogeneity differences of the disease.

Chromosomal alterations, changes in DNA copy number, tumor-promoting secondary somatic mutations and sequence mutations that drive leukemogenesis have become important hallmarks in the classification, pathogenesis and biomarker identification of Acute lymphoblastic leukemia based on comprehensive genetic studies including whole exome/genome sequencing, transcriptome analysis and genomic microarrays.

The majority of the genes involved in Acute lymphoblastic leukemia pathogenesis include transcriptional regulators, lymphoid signaling molecules and tumor suppressor genes related to lymphoid development. (Table 1.1.1). For instance, mutated PAX5 and IKZF1 genes drive progressive development of B-ALL (Table 1.1.2). Tumor suppressor genes like RB1 and CDKN2A/CDKN2B are altered by deletions and translocations in B-ALL.

Acute lymphoblastic leukemia is divided into two main categories, B-cell lymphoblastic leukemia/lymphoma and T-cell lymphoblastic leukemia/lymphomas based on Health Organization (WHO) revision in 2016.

B-cell lymphoblastic leukemia

B-cell lymphoblastic leukemia based on t (9;22) (q34; q11.2) [BCR-ABL1]

B-cell lymphoblastic leukemia based on t(v;11q23) [MLL rearranged]

B-cell lymphoblastic leukemia based on t (12;21) (p13; q22) [ETV6-RUNX1]

B-cell lymphoblastic leukemia based on t (1;19) (q23; p13.3) [TCF3-PBX1]

B-cell lymphoblastic leukemia based on t (5;14) (q31; q32) [IL3-IGH]

B-cell lymphoblastic leukemia based on intrachromosomic amplification of chromosome 21 (iAMP21)

B-cell lymphoblastic leukemia based on translocations related tyrosine kinases or cytokine receptors (Ph+ like Acute lymphoblastic leukemia)

B-cell lymphoblastic leukemia based on hyperdiploidy

B-cell lymphoblastic leukemia based on hypodiploidy

T-cell lymphoblastic leukemia

Early T-cell precursor lymphoblastic leukemia

B-ALL accounts for 75-80% while T-ALL accounts for 20-25% of all Acute lymphoblastic leukemia types. The subtypes of B-ALL have been classified based on recurrent genetic abnormalities including hypodiploidy, hyperdiploidy, t (9; 22) (BCR-ABL1), T (v; 11q23) (MLL rearranged) and BCR-ABL1-like (Ph like) Acute lymphoblastic leukemia. These cytogenetic abnormalities are related to the diagnosis status of both pediatric and adult B-ALL . For instance, Ph + ALL is known as the most common subtype of B-ALL cases (20-25% of all B-ALL) indicated in this classification with poor prognosis.

T-ALL is commonly observed at older age with dominancy in male sex and has poorer outcomes compared to B-ALL. T-ALL is characterized by mutations and deletions in the PHF6 tumor suppressor gene, which accounts for 16% of all T-ALL cases in children and 38% in adults. In addition to PHF6 gene mutation, activating NOTCH1 mutations, LMO2, MYB, WT1 and PTEN gene mutations, rearrangements of transcription factors like TLX1, LYL1, TAL1 and MLL were also observed in T-ALL.

Early T-cell precursor Acute lymphoblastic leukemia (ETP-ALL) is a subtype of T-ALLs distinguished by different cell surface markers with poor prognosis. These cells do not have CD1a and CD8 expressions while having weak CD5 expression and one or more myeloid-associated or stem cells associated markers.

Acute lymphoblastic leukemia is a treatable disease, especially in the pediatric population with a success rate of around 90%. However, this rate is around 30-35% in adults despite following the same treatment approaches. This is due to the fact that adults are more intolerant and resistant to chemotherapy and having risky genetic subtypes, mutations and epigenetic changes frequently.

Chemotherapy has been used as the standard cure for all Acute lymphoblastic leukemia types with different phases including the steroid pre-phase, the induction therapy, the consolidation and maintenance phases and central nervous system (CNS) prophylaxes. In different clinical setups, different combinations of drugs with different mechanisms of action have been used to remove the majority of the malignant Acute lymphoblastic leukemia cells and prevent drug resistance .

In the steroid pre-phase therapy, corticosteroids are used. Moreover, genetic and prognostic characterization of the disease such as the presence of RAS and CREBBP mutations might affect the therapy in the steroid pre-phase. In induction therapy which is the first stage of Acute lymphoblastic leukemia treatment, vincristine, methotrexate (MTX), anthracyclines including doxorubicin, daunorubucin, cytarabine are commonly used to provide normal blood cell production. Although this therapy provides a high rate of complete remission (CR), it causes severe side effects in children.

At the end of induction therapy, consolidation and maintanance therapy is given and eliminates residual leukemia cells. Various combinations of cytotoxic drugs used in the induction therapy are administered at high doses (like MTX and cytrabine). Hyper-CVAD (hyperfractionated Cyclophosphamide, Vincristine, Doxorubicin and Dexamethasone) is one of the most used protocols in this phase. After consolidation therapy, maintenance therapy lasts for 1-2 years and daily 6-mercaptopurine (6-MP) and weekly methotrexate (MTX) are given to patients.

Maintenance therapies are also strengthened by combining vincristine and steroids. Although maintenance therapy can provide CR, some obstacles including infection may result in death. In addition, a study showed that long-term maintenance therapy and high doses of 6-MP led to the development of secondary malignancies. Since chemotherapy-resistant cells might still remain following chemotherapy, allogeneic stem cell (allo-SCT) transplantation plays a significant role in eradicating remaining resistant cells. For patients at the relapse phase, especially for children, allogeneic stem cell (allo-SCT) transplantation is the backbone of the consolidation therapy.

However, complications such as infertility, growth retardation, metabolic diseases and secondary malignant neoplasms may occur after transplantation. Therefore, allo-SCT should be introduced to patients in the high-risk group if possible. The purpose of CNS prophylaxis is to prevent CNS relapse of the disease. Two main protocols which are intrathecal injection or high intravenous dose of MTX or Cytarabine (also used intrathecal, usually with steroids) are use are used to overcome the blood-brain barrier: d in order to overcome the blood-brain barrier.

CNS irradiation might be another option both for ALL adults and childhood. With this CNS relapse rate could be reduced. In addition to conventional chemotherapeutic approach and AlloSCT, targeted therapies such as immunotherapy, signaling pathway inhibition and CAR-T cell therapy have been revolutionized the therapy in ALL in favor of personalized medicine.

Philadelphia Positive Acute Lymphoblastic Leukemia

Ph + ALL is characterized by an unbalanced translocation between ABL gene located on chromosome 9 and BCR gene located on chromosome 22 which results in the formation of BCR-ABL fusion gene with abnormal tyrosine kinase (TK) activity. BCR-ABL fusion gene is predominantly found in chronic myeloid leukemia (CML), however, the presence of BCR-ABL is also a major pathogenicity factor in B-ALL (25%).

Newly formed BCR-ABL gene is responsible for the malignant transformation of the cells. BCR-ABL with different molecular sizes (p190, p210, p230) can be produced due to the different breakpoint regions in the BCR and ABL gene. Translocation commonly occurs between exons 1, 13/14 or exon 19 of BCR and a 140-kb region of ABL1 between exon 1b and 2 in all BCR-ABL positive hematological cancers such as CML, ALL and some AML cases.

The majority of the Ph+ ALL cases possess p190 kda protein, however, p210 kda protein can be also detected occasionally [28]. Exon 1 of BCR and exon 2 of ABL are fused to produce p190 BCR-ABL in ALL, which is also called minor breakpoint BCR (m-BCR).

Ph + Acute lymphoblastic leukemia Therapy

The incidence of Ph+ ALL increases with age reaching around 50% in patients above 60 years. Historically, intensive chemotherapy adapted from pediatric Acute lymphoblastic leukemia protocols was given to adult patients as a sole therapy which led to very poor outcomes such as short remission period and lower overall survival (OS) (<20%). Although CR was observed in patients receiving intensive chemotherapy, relapse was a major challenge for the patients who died within 6- 11 months after treatment.

Therefore, allogeneic stem cell transplantation (Allo-SCT) was thought as an effective treatment method in adult patients in the presence of suitable matched donors with increased OS (35-55%). If chemotherapy and AlloSCT were given together, significant success was observed with improved CR rates. However, finding available matched donors and decision for the number of AlloSCT trials represent major limitations.

The understanding of the molecular mechanism of Ph+ ALL and the role of BCR-ABL oncoprotein in leukemogenesis has opened the way of using TKIs which target TK activity of BCR-ABL. In addition to chemotherapy, AlloSCT, TKIs and combinational therapies, chimeric antigen receptor-modified (CAR) T cell therapy makes treatment approaches modernized. Moreover, the development of unique agents including inotuzumab (CD22 monoclonal antibody conjugated to the cytotoxic antibiotic calicheamicin) and blinatumomab (bispecific T cell engager anti-CD3 and CD19 antibody construct, resulted in a serial lysis of B cells by redirecting CD3+ T cells toward CD19+ B-ALL cells) have beneficial potentials for clinical usage especially for relapsed and/or refractory (R/R) Ph+ B-ALL.

Selective TKIs

The introduction of the first generation TKI, imatinib, which targets ATP binding domain of BCR-ABL to block its TK activity, into adult Ph+ ALL therapy had modest and unstable results, however, its combination with standard chemotherapy was safe and resulted in CR rates between 91% and 98% and OS rate reaching up to 50% (Treatment of Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia). Imatinib has been given to the patients together with chemotherapy by two general approaches which are simultaneous and successive methods.

Even though these combination therapies have antileukemic effects, the development of imatinib resistance throughout the therapy stays as a major problem. This resistance is divided into BCR-ABL dependent and independent mechanisms. BCR-ABL dependent mechanism is associated with the multiplication of BCR-ABL gene and point mutations in BCR-ABL that might be mutations at threonine 315 [T315] [57-59] and phenylalanine 317 [F317], at the Src homology 2 (SH2) binding site and at the ATP-binding pocket (in SH1 domain).

The most common mutation leading to conversion of glutamic acid to lysine at codon 255 (E255K) occurs principally after imatinib administration. Some mutations, particularly ATP-binding pocket mutations, are more resistant to imatinib and patients having these mutations are generally described with worse prognosis. BCR-ABL independent mechanisms are related to failure in drug uptake and efflux, altered alternative signaling pathways that promote abnormal cell proliferation and survival. For instance, the presence of BCR-ABL might increase multidrug resistance protein (MDR, PgP) expression which pumps imatinib out, therefore, intracellular concentration of imatinib is decreased.

The presence of secondary resistance to imatinib has resulted in the development of a number of second-and third-generation TKIs to overcome the resistance. Most common second generation TKIs are nilotinib and dasatinib. Nilotinib shows higher binding affinity for BCR-ABL and greater activity compared to imatinib. Moreover, it overcomes resistance to mutations that imatinib causes. In clinical studies from independent centers, nilotinib has been administrated to the patients in combination with chemotherapy and showed promising results with increased CR and OS rates.

Dasatinib, a second-generation TKI, inhibits both active and inactive forms of BCR-ABL and is 325-fold more effective than imatinib. It overcomes most of the imatinib-resistant kinase domain mutations. Dasatinib introduced into Ph+ ALL patients with resistance or intolerance to imatinib as single agent and showed some initial activities. In a study, 7 out of 10 patients treated with dasatinib achieved CR and 8 patients showed significant cytogenetic response.

In another study, 78% of 46 patients having BCR-ABL positive kinase domain mutations and patients (20%) carrying T315I showed remarkable hematologic and cytogenetic responses after dasatinib treatment. However, observed results were short-lived and progression free survival was around maximum 3 months. Therefore, combination of dasatinib with chemotherapeutic regimens have been investigated in various clinical studies. In a study, combination of dasatinib with chemotherapy hyper-CVAD: in 35 Ph+ ALL patients resulted in 94% CR and extended life span up to 2 year.

Patients, who relapse after therapy with imatinib, often develop kinase domain mutations responsible for imatinib resistance. T315I mutation, responsible for up to 75% of cases of acquired kinase mutations at the time of relapse, is known to be less sensitive to all first- and second-generation TKIs.Therefore, third generation TKIs such as ponatinib have been specifically designed to overcome most of the kinase domain mutations, including T315I.

Ponatinib introduced into Ph + ALL patients as single agent or in combination with chemotherapy. However, ponatinib in combination with intensive chemotherapy has shown the highest anti-leukemia activity. For example, ponatinib in combination with hyper-CVAD in Ph+ ALL was more effective than dasatinib in combination with hyper-CVAD (reference). Ponatinib did not only overcome T3151 mutation, but also resulted in higher CR. Currently, the standard treatment protocol includes the combination of a TKI with chemotherapy or corticosteroids. The major problem in combination setups due to the lack of randomized trials evaluating the advantage of one TKI over the others is which TKI should be preferred.