**5. Other factors potentially influencing resistance to myeloma therapies**

#### **5.1. Cytogenetics, mutation patterns and clonal evolution**

Cytogenetic abnormalities in MM are broadly divided into copy number changes or translocations, most commonly involving the immunoglobulin heavy chain gene [125]. Various cytogenetic abnormalities were shown to be associated with the likelihood of durable responses to therapy but they do not directly explain mechanisms of drug resistance or disease progression [126]. High risk genetic features frequently result in the dysregulation of transcription factors or tumour suppressors and include t(4;14), t(14;16), t(16;20), del(17p) and copy number changes of chromosome 1, which are used for stratifying MM patients in clinical trials and are now becoming important in guiding therapy in routine practice [126]. For example, the EMN02/HO95 study demonstrated the benefit of double autologous stem cell transplantation in patient with high-risk genetics, essentially negating the adverse prognosis of high genetic risk MM [127]. Similarly, the addition of Bortezomib to induction regimens in patients receiving HDM/ASCT may partially overcome cytogenetically defined poor risk [128]. On the other hand, patients with trisomies may respond particularly well to lenalidomide based protocols [129]. Mutational events such as those involving p53 are associated with particularly poor PFS, however, the significant heterogeneity of point mutational events elucidated in whole exome sequencing studies means generalisations of such molecular changes are not possible [130].

as to whether the MMSC derives from a clonotypic B cell (CD19<sup>+</sup>

robust MM reconstitution in the absence of a CD19<sup>+</sup>

between undifferentiated pre-plasma cells (CD19−

onciles inconsistencies surrounding the MMSC phenotype.

dehydrogenase (ALDH) have been demonstrated in CD138−

counterparts rendering the CD138−

CD138−

cell (CD19−

(CD19−

CD138<sup>+</sup>

cells and CD19−

**6. Conclusion**

CD138<sup>+</sup>

CD138−

Resistance Mechanisms to Novel Therapies in Myeloma http://dx.doi.org/10.5772/intechopen.77004

population [136]. To shed some light on

) and differentiated plasma cells

plasma cells compared to their

CD138<sup>+</sup>

plasma

population more resistant to certain chemothera-

pre-plasma cells harbour MMSC activity but exhibit differential resis-

CD138+/−). Clonotypic B cells were found to be resistant to a range of anti-MM

CD138−

) thus representing reversible, bi-directional phenotypic and functional states

therapies including Bortezomib and Lenalidomide and possessed a high drug efflux capacity [135]. However, clonotypic non-B cells have also been shown in many studies to result in

this dichotomy with respect to clonotypic non-B cells, there appears to be an interconversion

that share MMSC activity [137]. Furthermore, the pre-plasma cells were found to be more quiescent, primarily located at extramedullary sites, and up to 300-fold more drug resistant to agents including Bortezomib [137]. These informative findings imply phenotypic and functional plasticity between undifferentiated and differentiated clonotypic plasma cells which could explain why differentiated MM plasma cells possess clonogenic capacity and also rec-

Several factors have been attributed to the MMSC that confer drug resistance. (1) Side population (SP) MM cells, which possess stem-like properties, show stronger activity of several ABC transporters when compared to main population (MP) cells [138]. (2) High levels of aldehyde

peutic agents which result in the generation of toxic aldehyde intermediates that are metabolised by ALDH1 [135]. In one study, forced expression of member A1 of the ALDH1 family of proteins resulted in resistance to Bortezomib [139]. (3) Increased expression of Bcl-2 family members in MMSCs expressing the retinoid acid receptor alpha 2 (RARα2) endowed these cells with increased drug resistance [140], and more recently, increased expression of Bruton's

tance to treatment since pre-plasma cells are more quiescent than plasma cells, shown by a lower proportion of these cells in S phase of the cell cycle [137]. Finally, (5) the Wingless (Wnt), Hedgehog and Notch signalling pathways are all highly active in MMSCs and may be responsible for maintaining stem cell properties, propagating MM and promoting therapeutic

Continued improvements in the efficacy and toxicity profiles of an ever-expanding number of novel MM therapies are challenging the current paradigm of high-dose therapy and autologous stem cell transplantation for newly-diagnosed MM. However, despite these advances, resistance to novel agents has been observed and will continue to be observed, requiring innovative ways to circumvent this problem. Changing therapy from one novel agent containing treatment regimen to a different one upon MM progression or relapse is reasonable, however, there is often little scientific basis for choosing the sequence of such regimens and the era of precision medicine for MM patients remains distant. Moreover, the inability to tailor treatment

tyrosine kinase (BTK) in MMSCs also induced drug resistance [141]. (4) CD19−

resistance together with a supportive and protective BMME [142].

) or clonotypic non-B

83

The development of whole exome sequencing and copy number profiling was combined with cytogenetics in a landmark paper by a consortium of European and American groups [131]. This elegant paper demonstrated that the majority of MM patients had multiple sub-clones present at the time of diagnosis and that within sub-clones there could be differing mutational events potentially driving behaviour [131]. When serial MM samples were analysed, diverse patterns of clonal evolution were detected. In some cases, simple clonal selection could be observed following a linear pattern of clonal evolution [131]. Differential clonal responses could explain the clinical observation that a MM patient may respond to a treatment initially, lose this response, respond to another treatment and at the time of subsequent relapse respond again to the initial therapy [132]. Branching evolution was also observed in some progressing patients [131]. During disease evolution differing processes may contribute to the mutational repertoire and the relative contributions may vary over time in the same patient resulting in mutational heterogeneity, frequently with very few recurrent genes [131].

#### **5.2. The myeloma stem cell**

Identification of the multiple myeloma stem cell (MMSC) has been a challenge predominantly because an agreed phenotype with MM propagating potential has not been definitively established, in part due to differences in experimental techniques and assays. The dominant viewpoint is that clonotypic CD138− cells represent MMSCs, however, some researchers have also shown that clonotypic CD138<sup>+</sup> plasma cells have properties of cancer stem cells such as selfrenewal, tumour-initiating potential and drug resistance [133, 134]. Controversy also exists as to whether the MMSC derives from a clonotypic B cell (CD19<sup>+</sup> CD138− ) or clonotypic non-B cell (CD19− CD138+/−). Clonotypic B cells were found to be resistant to a range of anti-MM therapies including Bortezomib and Lenalidomide and possessed a high drug efflux capacity [135]. However, clonotypic non-B cells have also been shown in many studies to result in robust MM reconstitution in the absence of a CD19<sup>+</sup> population [136]. To shed some light on this dichotomy with respect to clonotypic non-B cells, there appears to be an interconversion between undifferentiated pre-plasma cells (CD19− CD138− ) and differentiated plasma cells (CD19− CD138<sup>+</sup> ) thus representing reversible, bi-directional phenotypic and functional states that share MMSC activity [137]. Furthermore, the pre-plasma cells were found to be more quiescent, primarily located at extramedullary sites, and up to 300-fold more drug resistant to agents including Bortezomib [137]. These informative findings imply phenotypic and functional plasticity between undifferentiated and differentiated clonotypic plasma cells which could explain why differentiated MM plasma cells possess clonogenic capacity and also reconciles inconsistencies surrounding the MMSC phenotype.

Several factors have been attributed to the MMSC that confer drug resistance. (1) Side population (SP) MM cells, which possess stem-like properties, show stronger activity of several ABC transporters when compared to main population (MP) cells [138]. (2) High levels of aldehyde dehydrogenase (ALDH) have been demonstrated in CD138− plasma cells compared to their CD138<sup>+</sup> counterparts rendering the CD138− population more resistant to certain chemotherapeutic agents which result in the generation of toxic aldehyde intermediates that are metabolised by ALDH1 [135]. In one study, forced expression of member A1 of the ALDH1 family of proteins resulted in resistance to Bortezomib [139]. (3) Increased expression of Bcl-2 family members in MMSCs expressing the retinoid acid receptor alpha 2 (RARα2) endowed these cells with increased drug resistance [140], and more recently, increased expression of Bruton's tyrosine kinase (BTK) in MMSCs also induced drug resistance [141]. (4) CD19− CD138<sup>+</sup> plasma cells and CD19− CD138− pre-plasma cells harbour MMSC activity but exhibit differential resistance to treatment since pre-plasma cells are more quiescent than plasma cells, shown by a lower proportion of these cells in S phase of the cell cycle [137]. Finally, (5) the Wingless (Wnt), Hedgehog and Notch signalling pathways are all highly active in MMSCs and may be responsible for maintaining stem cell properties, propagating MM and promoting therapeutic resistance together with a supportive and protective BMME [142].
