Myeloma cells are highly dependent on the unfolded protein response to assemble folded immunoglobulins correctly. Therefore, targeting protein handling within a myeloma cell by inhibiting the aminopeptidase enzyme system, which catalyses the hydrolysis of amino acids from the proteins NH 2 terminus, represents a therapeutic approach.

Blastoid morphology is a rare presenting feature of myeloma which is frequently seen in patients with extramedullary myeloma and is associated with poor clinical outcome. Cell cycle active agents can be effective as treatment for aggressive myeloma and their activity enhanced by using them in combination with the anti-angiogenic agent thalidomide. DT-PACE is an example of such a regimen which we have used to treat 26 relapsed and or refractory patients with extramedullary/blastoid myeloma. The overall response rate (complete response/PR) was 59%, but despite these initial good responses, patients had a short progression free survival (PFS) and overall survival (OS). A subgroup of patients who proceeded to autologous stem cell transplant (ASCT) have a trend towards a better PFS and OS when compared with the group receiving chemotherapy alone (PFS = 10 vs. 3 months P = 0.273 and OS 10 vs. 7 months P = 0.235). Interestingly of the group who received ASCT consolidation three patients remain alive beyond 18 months. In conclusion, the clinical outcome of this group of cases is poor even when treated with the intensive regimen DT-PACE; however, a subgroup can do well if DT-PACE is consolidated by ASCT.

A 64-year-old Caucasian man was diagnosed with multiple myeloma in 1998, following radiotherapy for a solitary sternal plasmacytoma 12 months previously. He received nine monthly courses of oral melphalan followed by six courses of oral dexamethasone. In August 1999 he was commenced on oral cyclophosphamide 100 mg ⁄ d, and reported the development of pigmented nails of hands and feet some 5 months later. Cyclophosphamide therapy is documented to rarely cause black pigmentation of the nails after a total dose of between 1AE2 and 12AE3 g. These changes start in the proximal nail beds and progress distally; on withdrawal of cyclophosphamide, resolution proceeds in a similar fashion. The mechanism of nail pigmentation remains unknown, although the initial involvement of the nail bed, the band pattern and distal migration of the band with subsequent disappearance suggests a disturbance of the nail plate.

Multiple myeloma is the second most common haematological malignancy. It is becoming increasingly manageable with conventional and high-dose chemotherapy but there remains a critical need to develop both new drugs and combinations to improve long-term outcomes. Novel biological therapies that specifically target myeloma cells and/or their microenvironmental interactions are being developed that are highly effective, both as single agents and as combinations. Chief among these new agents are the proteasome inhibitor, bortezomib, and the immunomodulatory agents, thalidomide and lenalidomide. These drugs show improved single agent activity that is enhanced in combination. However, many drugs that are being developed in this setting may only have limited single agent activity, but combination use with these and other agents represents a very exciting way of targeting important pathogenic pathways crucial in myeloma development. This represents a challenge for both drug development and clinical trial evaluation, which has the potential to revolutionise the clinical management of myeloma and a paradigm for drug development in other diseases.

4083 In normal cells, autophagy is up-regulated under conditions of stress ensuring cell survival, suggesting that if it is inhibited cell death will ensue. In some cancer settings however it has been shown to enhance cell death. Therefore if autophagy is to be a target for anti-cancer therapy, it is important to determine whether to promote or inhibit the process. Cellular entry to autophagy can be manipulated via the PI3K/AKT/mTOR pathway, a pathway known to be important for myeloma cell growth and survival. In this study we investigated the effects of PI-103, a dual Class I PI3K and mTOR inhibitor, which activates autophagy, and used it as a tool to investigate the interaction of autophagy with other myeloma therapies. As the PI3K/AKT/mTOR pathway has been suggested to be a central pathway controlling entry into autophagy, we first determined the basal expression of key members of the pathway in a panel of myeloma cell lines. PI3K alpha, beta delta and gamma isoforms were expressed to varying degrees and constitutive activation of the pathway, (pAKT or pMTOR), was seen in the majority of cell lines. As predicted, treatment with PI-103 induced autophagy in myeloma cells as demonstrated by an increase in cellular inclusions staining positively with acridine orange, cleavage of the autophagosome marker, LC3, and a decrease in p62. PI-103 was shown to inhibit proliferation of all the myeloma cell lines and patient cells tested to varying degrees, although 100% growth inhibition was not seen. Bone marrow stromal cells were unaffected. The main mode of action of PI-103 was autophagy activation, and in keeping with this the extent of cell death measured by Annexin V/PI binding and trypan blue exclusion, was minimal. Cell cycle analysis demonstrated an increase in G0-G1 phase. The unfolded protein response (UPR) is important in myeloma cells and UPR activation and autophagy have been reported to be interlinked. Following exposure to PI-103, splicing of XBP1 mRNA to its active form, XBP1s, was seen, CHOP and ATF4 mRNA levels were also increased, consistent with activation of at least two branches of the UPR in response to PI3K/mTOR inhibition and autophagy induction. As myeloma cells activate autophagy as a pro-survival pathway following PI3-kinase inhibition, we were interested to understand the effect of blocking autophagy in this context. When PI-103 was combined with the autophagy inhibitor, Bafilomycin greatly enhanced apoptosis was seen. This increased apoptosis was seen in cells constitutively expressing p-AKT, with a complete loss of both phospho- and total levels of AKT and mTOR, an increase in the cleaved forms of caspase 3 and Bcl2, and massive activation of the IRE1 and PERK branches of the UPR. This effect was not seen in cells lacking p-AKT, a phenomenon described as ‘context-dependent oncogene addiction' suggesting that measurement of p-AKT may be a useful predictive marker for response to joint PI3K/autophagy inhibition. Importantly the pro-apoptotic effects of the combination are not overcome by the presence of bone marrow cytokines, a more representative model of the physiological situation in vivo. In conclusion our data highlights the interplay between known myeloma growth and survival pathways and autophagy and suggests that combining PI3K inhibitors, with agents that target autophagy, may be beneficial for the treatment of myeloma, particularly in >50% of patients that express p-AKT. Disclosures: No relevant conflicts of interest to declare.