AKT Dependent and AKT Independent Multiple Myeloma: Heterogeneity in Tumor Cell Signaling.

Abstract The PI3K/Akt pathway has been reported to critically contribute to survival and malignant growth of multiple myeloma (MM). Because most of these data are based on pharmacologic inhibition it is not clear if the effects are due to Akt inhibition or off-target effects. Furthermore, the Akt family of kinases consists of three highly homologous isoforms, that may, nonetheless, display individual functional properties. We therefore conducted siRNA experiments to evaluate if any single isoform posesses a special role for the viability of MM cells. This was complemented with extensive analyses into the functional and signaling properties of the Akt pathway in primary MM cells (n = 30). Our knock-down experiments revealed that in MM.1S, an MM cell line with constitutive phospho-Akt signaling, Akt1 and Akt2 both contributed to MM cell survival whereas Akt3 seemed to be of less relevance. Conversely, survival of MM cell line AMO-1 which has no constitutive phospho-Akt signal was completely unaffected. Treatment of these MM cell lines with the Akt1 and Akt2 specific inhibitor Akti-1/2 showed that this drug totally abolished the phospho-Akt signal in MM.1S at a concentration of 10 microM. Again, MM.1S cells underwent apoptosis whereas AMO-1 cells were resistant. Next, we analyzed Akt signaling in a large panel of primary MM samples. Phosphorylated Akt was determined with intracellular staining and flow cytometry analysis in primary tumor samples and could be detected in about 60% of MM cases. This constitutive signal could be blocked with Akti-1/2 in the presence and absence of bone marrow stromal cells. Pharmacologic inhibition of Akt led to strong induction of cell death in 46% of primary MM samples, whereas the rest was largely resistant to Akt inhibition. The samples sensitive to Akt inhibition were mostly identical to those that displayed a constitutive phospho-Akt signal. Of interest, there was no correlation between Akt dependence and mutational inactivation of PTEN. Further inhibition of other signaling cascades implicated in growth and survival of MM cells, such as the MAPK or STAT3 pathways, had only minor additional effects on tumor cell viability of samples resistant to Akt inhibition. Our analysis indicates substantial heterogeneity in MM cells that defines Akt dependent and Akt independent MM subgroups. Akt1 and Akt2 proved relevant for the survival of subsets of MM cell lines and primary samples. Taken together, this is the first comprehensive functional and molecular signaling analysis of primary MM samples which led to the identification of novel functionally defined myeloma subgroups.