Bone Marrow Aging and the Leukaemia-Induced Senescence of Mesenchymal Stem/Stromal Cells: Exploring Similarities.

Bone marrow aging is associated with multiple cellular dysfunctions, including perturbed haematopoiesis, the propensity to haematological transformation, and the maintenance of leukaemia. It has been shown that instructive signals from different leukemic cells are delivered to stromal cells to remodel the bone marrow into a supportive leukemic niche. In particular, cellular senescence, a physiological program with both beneficial and deleterious effects on the health of the organisms, may be responsible for the increased incidence of haematological malignancies in the elderly and for the survival of diverse leukemic cells. Here, we will review the connection between BM aging and cellular senescence and the role that these processes play in leukaemia progression. Specifically, we discuss the role of mesenchymal stem cells as a central component of the supportive niche. Due to the specificity of the genetic defects present in leukaemia, one would think that bone marrow alterations would also have particular changes, making it difficult to envisage a shared therapeutic use. We have tried to summarize the coincident features present in BM stromal cells during aging and senescence and in two different leukaemias, acute myeloid leukaemia, with high frequency in the elderly, and B-acute lymphoblastic leukaemia, mainly a childhood disease. We propose that mesenchymal stem cells are similarly affected in these different leukaemias, and that the changes that we observed in terms of cellular function, redox balance, genetics and epigenetics, soluble factor repertoire and stemness are equivalent to those occurring during BM aging and cellular senescence. These coincident features may be used to explore strategies useful to treat various haematological malignancies.

Sensitization to Drug Treatment in Precursor B-Cell Acute Lymphoblastic Leukemia Is Not Achieved by Stromal NF-κB Inhibition of Cell Adhesion but by Stromal PKC-Dependent Inhibition of ABC Transporters Activity.

Cell adhesion to stromal support and the associated intracellular signaling are central to drug resistance, therefore blocking both has been effective in increasing drug sensitization in leukemia. The stromal Ser/Thr protein kinase C (PKC) has been found to be important for conferring protection to leukemic cells. We aimed at elucidating the intracellular signals connected to cell adhesion and to stromal PKC. We found that NF-κB and Akt were up-regulated in mesenchymal stem cells (MSC) after binding of B-cell acute lymphoblastic leukemia (B-ALL) cells. Nevertheless, Akt inhibition did not induce B-ALL cell detachment. In spite of a clear activation of the NF-κB signaling pathway after B-ALL cell binding (up-regulation NF-κB1/2, and down-regulation of the IKBε and IKBα inhibitors) and an important reduction in cell adhesion after NF-κB inhibition, sensitization to the drug treatment was not observed. This was opposite to the PKC inhibitors Enzastaurin and HKPS, a novel chimeric peptide inhibitor, that were able to increase sensitization to dexamethasone, methotrexate, and vincristine. PLCγ1, Erk1/2, and CREB appear to be related to PKC signaling and PKC effect on drug sensitization since they were contra-regulated by HKPS when compared to dexamethasone-treated cells. Additionally, PKC inhibition by HKPS, but not by Enzastaurin, in MSC reduced the activity of three ABC transporters in leukemic cells treated with dexamethasone, a new indirect mechanism to increase sensitization to drug treatment in B-ALL cells. Our results show the validity of targeting the functional characteristic acquired and modulated during cell-to-cell interactions occurring in the leukemic niche.

Leukemia-Induced Cellular Senescence and Stemness Alterations in Mesenchymal Stem Cells Are Reversible upon Withdrawal of B-Cell Acute Lymphoblastic Leukemia Cells.

Leukemic cell growth in the bone marrow (BM) induces a very stressful condition. Mesenchymal stem cells (MSC), a key component of this BM niche, are affected in several ways with unfavorable consequences on hematopoietic stem cells favoring leukemic cells. These alterations in MSC during B-cell acute lymphoblastic leukemia (B-ALL) have not been fully studied. In this work, we have compared the modifications that occur in an in vitro leukemic niche (LN) with those observed in MSC isolated from B-ALL patients. MSC in this LN niche showed features of a senescence process, i.e., altered morphology, increased senescence-associated ß-Galactosidase (SA-ßGAL) activity, and upregulation of p53 and p21 (without p16 expression), cell-cycle arrest, reduced clonogenicity, and some moderated changes in stemness properties. Importantly, almost all of these features were found in MSC isolated from B-ALL patients. These alterations rendered B-ALL cells susceptible to the chemotherapeutic agent dexamethasone. The senescent process seems to be transient since when leukemic cells are removed, normal MSC morphology is re-established, SA-ßGAL expression is diminished, and MSC are capable of re-entering cell cycle. In addition, few cells showed low γH2AX phosphorylation that was reduced to basal levels upon cultivation. The reversibility of the senescent process in MSC must impinge important biological and clinical significance depending on cell interactions in the bone marrow at different stages of disease progression in B-ALL.

Dual Targeting of Stromal Cell Support and Leukemic Cell Growth by a Peptidic PKC Inhibitor Shows Effectiveness against B-ALL.

Mesenchymal stem cells (MSC) favour a scenario where leukemic cells survive. The protein kinase C (PKC) is essential to confer MSC support to leukemic cells and may be responsible for the intrinsic leukemic cell growth. Here we have evaluated the capacity of a chimeric peptide (HKPS), directed against classical PKC isoforms, to inhibit leukemic cell growth. HKPS was able to strongly inhibit viability of different leukemic cell lines, while control HK and PS peptides had no effect. Further testing showed that 30% of primary samples from paediatric B-cell acute lymphoblastic leukaemia (B-ALL) were also strongly affected by HKPS. We showed that HKPS disrupted the supportive effect of MSC that promote leukemic cell survival. Interestingly, ICAM-1 and VLA-5 expression increased in MSC during the co-cultures with B-ALL cells, and we found that HKPS inhibited the interaction between MSC and B-ALL cells due to a reduction in the expression of these adhesion molecules. Of note, the susceptibility of B-ALL cells to dexamethasone increased when MSC were treated with HKPS. These results show the relevance of these molecular interactions in the leukemic niche. The use of HKPS may be a new strategy to disrupt intercellular communications, increasing susceptibility to therapy, and at the same time, directly affecting the growth of PKC-dependent leukemic cells.