For an example, if a pathway consists of a cascade of reactions in such a way that phosphorylation is only required as its trigger, then such pathway would not be fully inhibited by SAR131675 activated PKC inhibitors. Namely, the transient PKC activity in the presence of activated PKC inhibitors would be sufficient to activate the pathway. This limited efficacy of active PKC inhibitors due to the lag time of inhibitor binding could be an alternative mechanism for resistance to kinase inhibitors in addition to protection through scaffold proteins. On the other hand, activated PKC inhibition would be beneficial for therapeutic purposes. Many pathogenic pathways involve constitutively activated kinases, while normal pathways remain quiescent until they are activated by physiological stimuli. Thus, activated kinase inhibitors would selectively target such pathological pathways. These state-dependent inhibitions would be a useful strategy to target selective conditions in signaling cascades. The incidence of melanoma has been constantly increasing during the last decades. Adjuvant therapy after complete resection is recommended for thick primary melanoma with lymph node metastases, because recurrence rates are relatively high and overall survival is poor. However, IFNalpha remains the only approved adjuvant therapy, which provides a modest disease-free survival benefit. Furthermore, it is especially GW-572016 concerning that the conventionally used drugs for metastatic melanoma include dacarbazine and IL-2, both of which cause poor and transient responses. Although promising therapeutic responses have been observed in recent clinical trials using the BRAF inhibitor Vemurafenib and the monoclonal antibody against CTLA-4 Ipilimumab, both recently approved by the FDA, emergence of resistance and severe side effects have already been confronted. Aberrant NOTCH signaling has also been shown to confer stem cell-like properties in different cancer types, such as breast cancer and glioma. Identification of stem cell-like tumor initiating cells has been of major interest in melanoma. Although there is an ongoing debate about the 
frequency and identity of melanoma initiating cells, the inability to eradicate this subpopulation is thought to be a reason for the failure of current treatment strategies. Therefore, NOTCH inhibition in melanoma, possibly through the targeting of tumor initiating cells, can be foreseen as a new and promising therapeutic strategy. Essential to NOTCH signaling is the catalytic cleavage of NOTCH receptor by the gamma secretase complex. Different inhibitors of gamma secretase have been developed. These inhibitors have been tested in vitro on a variety of cell lines, including melanoma. Clinical data have been supplied mostly by trials in adult T Cell leukemia, but efficacy has been hindered by significant gastrointestinal toxicities associated with treatment. However, RO4929097 is a novel gamma secretase inhibitor with an improved clinical toxicity profile. Here, we report the preclinical effects of RO4929097 on both primary and metastatic melanoma cells. In particular, we show for the first time that the inhibition of NOTCH signaling has an impact on the tumor initiating properties of melanoma cells. The aggressiveness of melanoma, which is surprisingly high considering that it is among a handful of cancers whose dimensions are reported in millimeters, is due to the high degree of heterogeneity and plasticity combined with the chemoresistance of melanoma cells.