**3. What is the consequence of the t(9;22)?**

The results of the chromosomal translocation occurring between the chromosome 9 and 22, t(9;22), are a longer chromosome 9 (9q+) and a smaller derivative chromosome 22, the Ph [7, 8]. By the eighties of the past century the molecular characterization of the Ph led to the identification of a novel chimeric gene, BCR-ABL, which later on has been found to encode for a chimeric protein with a constitutive tyrosine kinase activity and with potent oncogenic properties [9, 10]. The c-ABL and Breakpoint of Cluster Region (BCR) loci are localized on the long arm of the chromosome 9 and 22, respectively [11]. Depending on the different breakpoints occurring on the two chromosomes resulting in different BCR-ABL variants. Though, all BCR and c-ABL DNA breakpoints fall within intronic regions those occurring in the BCR gene are highly variable and thus responsible for defining the major differences among the different variants. The variation in the BCR part of the fusion transcript contrasts with the constant c-ABL part. Indeed, all the breakpoints so far identified within the c-ABL gene occur in a large (300-kb) region in the 5′ portion of the gene, localized upstream of the exon 2, and generally falling in the intron sequences restricted between the two alternative first exons (1b and 1a). Regardless the structure of the different fusion genes the BCR exons directly fuse to the second c-ABL exon (a2). The most frequent BCR-ABL fusion variant is the p210 in which the BCR exon 13, or 14, is fused downstream of the alternative exons 1 of the c-ABL gene and thus leading to a fusion protein with approximately the first half from BCR and the remaining second half from ABL. Mostly this variant is found in CML patients accounting for approximately 95% of the BCR-ABL fusion gene in all the CML cases. A second frequent variant, p190, is found in approximately 20–30% of adult patients with Acute Lymphoblast Leukemia (ALL) [12] and, very rarely, also in Acute Myeloid Leukemia (AML) [13]. When compared to the p210 variant, in this case the breakpoint within the BCR locus is localized in the 3′ half of the first BCR intron, thus encoding for a shorter BCR portion (approximately 425 aminoacids). The third most common BCR-ABL variant, p230, is the largest and is defined by a breakpoint cluster region encompassed between the exons 19 and 21. Whereas the p190 characterizes a more acute form of leukemia usually of lymphoid origin, the latter variant is peculiar of neutrophilic CML. Besides, there are additional BCR-ABL variants, though they have been observed less frequently. Interestingly, some of them are peculiar because they are the results of alternative splicing leading to truncated chimeric proteins that are all lacking tyrosine kinase activity [14]. Furthermore, in hematopoietic malignancies, the BCR gene has been identified

*Advances in Precision Medicine Oncology*

including acute promyelocytic leukemia.

cessfully cured in the vast majority of the cases.

containing part of normal chromosomes [4].

**chromosome?**

abnormality was a shortened chromosome 22. Among a chorus of skepticism and wonder at the beginning of the seventies that short chromosome, that it is now known as Philadelphia chromosome (Ph), was identified as the product of a reciprocal translocation between the long arms of chromosomes 9 and 22, t(9;22). However, we had to wait until the eighties to know that the exact molecular consequence of the t(9;22) was a fusion gene encoding for a chimeric protein displaying constitutively tyrosine kinase activity. Altogether these discoveries delivered an outstanding message whereby a disease was tightly linked to a single oncogene, BCR-ABL. Since then, dozens of translocations have been found in other cancers,

BCR-ABL is a peculiar protein for several of reasons: 1) it is a chimeric protein that is encoded by a fusion gene deriving from a reciprocal chromosomal translocation; 2) it is a constitutively active tyrosine kinase eliciting oncogenic signals, 3) it has been the first oncogene associated to a disease displaying dual properties either as driver and in sustaining the neoplasm evolution, and 4) it has been the first kinase to be selectively targeted with small molecules, thus paving the way for the

In the present chapter we are going to discuss the milestones of a story, started 60 years ago, which has happily led to the selective pharmacological inhibition of BCR-ABL. Hence, CML, whose diagnosis was before a death sentence, is now suc-

**2. Is there any specific reason underlying the generation of the Ph** 

Basically, every chromosomal translocations require DNA Double-strand breaks (DSBs) in two different locations and that the broken ends of non-

homologous chromosomes are fused together. DNA double-strands breaks might be due to different causes (*e.g.* ionizing radiation, reactive oxygen species, DNA replication across a nick, malfunctioning of DNA metabolic enzymes such as type II DNA topoisomerase or RAG complex during illegitimate V(D)J recombination). Cells to preserve their genome integrity upon DNA damage respond by activating a repair machinery that should catalyzes the joining of the broken ends [3]. However, the outcome of the joining process leads to a variety of rearrangement. For instance, precise joining of broken ends can generate a normal chromosome. Inversions, deletions and duplications can occur when joining involves two broken ends on the same chromosome. Non-Homologous End Joining (NHEJ) is often imprecise; thus some nucleotides may be lost during the joining process. Eventually, translocations may occur when the broken ends of two nonhomologous chromosomes are joined together thus leading to novel chromosomes

Aside these notions, currently our knowledge regarding the molecular mechanisms responsible for the reciprocal chromosomal translocation occurring between the chromosome 9 and the 22, t(9;22), generating the Philadelphia chromosome (Ph), remain still rather elusive. Fundamentally, it has been speculated that there are two plausible hypotheses. One view prefers to lean towards an entirely random "breaking and re-ligating process" occurring with relatively similar frequency between any two chromosomes within a cell. Chromosomal translocations that give and adaptive advantage are pretty rare and associated with negative consequences (*e.g*. cancer). The success of the t(9;22) can be explained by the fact that, the resulting fusion gene encodes for a protein with transforming properties conferring selective fitness advantages to the host cell. Conversely, by virtue of this, any

development of a number of tyrosine kinase inhibitors (TKIs).

**232**

fused to multiple tyrosine kinases encoding genes, other than ABL, including Fibroblast Growth Factor Receptor1 (FGFR1) -t(8;22)- [15, 16], Platelet Derived Growth Factor Receptor A (PDGFRA) -t(4;22)- [17, 18], RET -t(10;22)- [19] and Jak2 -t(9;22)-[20–22] producing different fusion transcripts that are all encoding for cytoplasmic chimeric proteins displaying dysregulated tyrosine kinase enzymatic activity and onocogenic properties. The causal reason behind the commonality of BCR as fusion partner is not well understood. As we have previously discussed it has been speculated that genes such as BCR are located near chromosomal fragile sites that show breaks or gaps on metaphase chromosomes due to replication stress which are prone to breakage and translocation as result. Interestingly, though BCR fusion genes have also been detected in solid tumors, to date BCR fusion proteins that behave as cancer drivers have solely been identified in hematological cancers.
