Leukemia & Lymphoma

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PI3K/AKT/mTOR pathway in multiple myeloma:

from basic biology to clinical promise

Vijay Ramakrishnan & Shaji Kumar

To cite this article: Vijay Ramakrishnan & Shaji Kumar (2018) PI3K/AKT/mTOR pathway in
multiple myeloma: from basic biology to clinical promise, Leukemia & Lymphoma, 59:11,
2524-2534, DOI: 10.1080/10428194.2017.1421760

To link to this article: https://doi.org/10.1080/10428194.2017.1421760

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LEUKEMIA & LYMPHOMA
2018, VOL. 59, NO. 11, 2524–2534
https://doi.org/10.1080/10428194.2017.1421760

REVIEW

PI3K/AKT/mTOR pathway in multiple myeloma: from basic biology to clinical

promise
Vijay Ramakrishnan and Shaji Kumar
Division of Hematology, Mayo Clinic, Rochester, MN, USA

ABSTRACT ARTICLE HISTORY

Multiple myeloma (MM), a cancer of terminally differentiated plasma cells, is the second most Received 27 October 2017
common hematological malignancy. The disease is characterized by the accumulation of abnor- Revised 6 December 2017
mal plasma cells in the bone marrow that remains in close association with other cells in the Accepted 9 December 2017
marrow microenvironment. In addition to the genomic alterations that commonly occur in MM,
KEYWORDS
the interaction with cells in the marrow microenvironment promotes signaling events within the Multiple myeloma; PI3K;
myeloma cells that enhances survival of MM cells. The phosphoinositide 3-kinase (PI3K)/protein AKT; mTOR; apoptosis;
kinase B (AKT)/mammalian target of rapamycin (mTOR) is such a pathway that is aberrantly acti- signaling
vated in a large proportion of MM patients through numerous mechanisms and can play a role
in resistance to several existing therapies making this a central pathway in MM pathophysiology.
Here, we review the pathway, its role in MM, promising preclinical results obtained thus far and
the clinical promise that drugs targeting this pathway have in MM.

Introduction malignancy of plasma cells and accounts for over

12,000 deaths per year just in the USA (https://seer.
Targeting abnormal signaling cascades specific to can-
cancer.gov/statfacts/html/mulmy.html). MM is always
cer cells continues to be an area of intense research.
preceded by premalignant conditions called
With a better understanding of these signaling events
Monoclonal Gammopathy of Undetermined
and the availability of potent and specific small mol-
Significance (MGUS) and Smoldering Multiple Myeloma
ecule inhibitors targeting such events, there is a strong
(SMM) [1–4]. Several studies have attempted to better
potential for more such agents to be incorporated into
understand the factor(s) that contribute to disease
the treatment armamentarium for various cancers.
transformation. The results from such studies have
Several such targeted agents have already shown clin-
educated us on the complex nature of MM, which
ical value such as vemurafenib in B-Raf proto-onco-
now is well recognized to be heterogeneous with a
gene, serine/threonine kinase (BRAF) mutated
multi-step evolution pattern. First, cytogenetic abnor-
melanoma, ceritinib in anaplastic lymphoma kinase
malities involving the Immunoglobulin heavy chain
(ALK) rearranged non-small cell lung cancer and imati-
locus are observed in MGUS, SMM, and active MM
nib in Philadelphia chromosome positive chronic mye-
patients. Some of the commonly observed transloca-
logenous leukemia (CML), and acute lymphoblastic
tions are t(11;14), t(4;14), t(14;16), and t(14;20) resulting
leukemia (ALL). Here, we review the central role played
in elevated levels of cyclin D, FGFR3, and MMSET,
by the PI3K/AKT/mTOR pathway in multiple myeloma
c-MAF, and MAFB, respectively. SMM patients having
(MM) growth and survival, promising preclinical results
these translocations are at a greater risk of progression
from studies examining inhibitors of this pathway, and
to active MM than those with no translocations. SMM
the potential for such drugs as therapeutic agents in
patients with t(4:14) have the highest risk with a
the management of MM.
median progression time of 28 months [5–9]. t(4;14) is
significantly more prevalent in active MM when com-
Multiple myeloma – a cancer with multiple
pared with premalignant conditions whereas t(14;16) is
genetic abnormalities
significantly more prevalent in patients with plasma
MM is a devastating disease that affects around 30,000 cell leukemia when compared with premalignant con-
people annually in the USA. It is an incurable ditions and MM [6,7,10]. Second, mutations in RAS,

CONTACT Vijay Ramakrishnan Ramakrishnan.vijay@mayo.edu Mayo Clinic, Division of Hematology, Mayo Clinic, 200 First St SW, Rochester, MN
55905, USA
ß 2018 Informa UK Limited, trading as Taylor & Francis Group

amplification of Myc, and mutations in the NF-jB path-
way are commonly observed in active MM but not in
MGUS and SMM and are thought to be driver muta-
tions and alterations for progression [11–15]. Third,
MM patients can acquire additional mutations in RAS,
p53, Cereblon (CRBN), IRF4, XBP1 with disease progres-
sion or response to therapies [16]. Thus, MM quite
expectedly has been a difficult disease to treat and
manage given the heterogeneity observed in patients.
In spite of this, MM has had a dramatic increase in sur-
vival post incorporation of immunomodulatory drugs
and proteasome inhibitors to the treatment regimen
[17]. However, patients continue to relapse and
become refractory to existing therapies. Hence, tar-
geted therapies based on mutational status or aber-
rant signaling events are areas of intense research in
MM.
In addition to these genetic alterations, MM plasma
cells are very dependent on the supporting cells in the
bone marrow microenvironment for their growth and
survival. The intimate association of the MM cells with
the stromal and endothelial cells in the microenviron-
ment contributes to elevated levels of cytokines and
cytokine stimulated pro-survival pathways such as the
PI3K/AKT/mTOR pathway within the plasma cells that
are vital for MM cells to grow, survive, and develop
resistance to existing therapies [18,19].
PI3K/AKT/mTOR – a pathway commonly
activated in MM
The PI3K/AKT/mTOR pathway has been extensively
studied and we would like to direct the readers to
excellent reviews published on the pathway and its
role in various malignancies [20–22]. In MM, activating
mutations in PI3K and AKT have not been identified
[23]. Similarly, deficiencies in phosphatase and tensin
homolog (PTEN) also occur in a very small proportion
of MM patients [24]. In spite of all this, the pathway is
activated and is of importance for MM cell survival
and growth. MM is predominantly a cancer localized
to the bone marrow where the cancerous plasma cells
remain in close direct and indirect communication
with other non-tumor cells in the bone marrow micro-
environment like stromal and endothelial cells. These
interactions lead to increased secretion of interleukin 6
(IL6) by the stromal cells and vascular endothelial
growth factor (VEGF) and insulin-like growth factor
(IGF) by MM cells. These cytokines activate their
respective receptors expressed on MM cells to up
regulate signaling events such as the PI3K/AKT/mTOR,
mitogen-activated protein kinase kinase/extracellular
signal-regulated kinase (MEK/ERK) and janus kinase/
AKT PATHWAY IN MYELOMA 2525
signal transducer and activator of transcription (Jak/
Stat) pathways and promote tumor growth [25]. The
complexity of the PI3K/AKT/mTOR pathway is ampli-
fied by the presence of numerous feedback loops that
exist within this pathway and the crosstalk with
numerous other pathways.
Feedback loops within the PI3K/AKT/mTOR
pathway
As stated above, IGF binds to IGF receptor (IGFR)
expressed on plasma cells, which then activates PI3K
through the phosphorylation of insulin receptor sub-
strate (IRS) 1 and 2 at tyrosine (Tyr) residues. Activated
PI3K then recruits pleckstrin homology (PH) domain
containing proteins 3-phospho inositide-dependent
protein kinase 1 (PDK1) and AKT to the cell surface,
where AKT gets phosphorylated at Threonine (Thr) 308
and activated by PDK1. Once AKT gets activated, it
promotes cell proliferation, anti-apoptosis, and metab-
olism signals leading to tumor growth and survival
[21,22,26,27]. A key downstream target of AKT is the
mammalian target of rapamycin (mTOR) complex 1
(mTORC1) that promotes protein synthesis, glycolysis
and inhibits autophagy (Figure 1). In addition, acti-
vated mTORC1 causes inhibitory phosphorylation of
IRS1 and 2 leading to feedback inhibition of the PI3K/
AKT pathway [20,26]. mTOR exists as two distinct multi
protein complexes namely mTORC1 and mTORC2. The
catalytic member of both these complexes is mTOR
with as many as five and six other binding partners in
mTORC1 and mTORC2, respectively [28–36]. While
mTORC1 is regulated by phospho (p) AKT (Thr 308), it
is now well known that mTORC2 is the kinase that
phosphorylates AKT at the serine (Ser) 473 residue
[20]. Once phosphorylated at the Ser 473 residue, AKT
modulates survival and cell cycle progression signals
through forkhead box protein O1 (FoxO1) phosphoryl-
ation and serum-and glucocorticoid-induced protein
kinase (SGK) activation [26]. Furthermore, it has been
suggested that the mTOR complexes cross inhibit each
other complicating the pathway activation status [37].
In addition to the above-mentioned feedback loops,
MM cells modulate the PI3K/AKT/mTOR pathway
through other unique mechanisms. Peterson et al.
identified an mTOR interacting protein DEP domain
containing TOR interacting protein (DEPTOR) that was
able to inhibit both mTORC1 and mTORC2 [33]. As
expected, DEPTOR was expressed at low levels in vari-
ous tumors. Surprisingly, DEPTOR was highly expressed
in MM cells. The authors found that overexpression of
DEPTOR inhibited mTORC1, which then relieved the
feedback inhibition of ribosomal protein S6 kinase-
2526 V. RAMAKRISHNAN AND S. KUMAR

Figure 1. Overview of the PI3K/AKT/mTOR pathway and the feedback loops that have been documented within this pathway in
MM. IGF1: insulin-like growth factor1; IGF1R: IGF1 receptor; VEGF: vascular endothelial growth factor; VEGFR: VEGF receptor;
PI3K: phosphoinositide 3-kinase; PDK: phosphoinositide-dependent kinase 1; AKT: protein kinase B; TSC1/2: tuberous sclerosis
protein 1/2; mTORC1: mammalian target of rapamycin complex 1; mTORC2: mammalian target of rapamycin complex 2; Raptor:
regulatory-associated protein of mTOR; DEPTOR: DEP domain containing TOR interacting protein; Rictor: rapamycin-insensitive com-
panion of mammalian target of rapamycin (RICTOR); p70S6K: ribosomal protein S6 kinase; 4EBP1: eukaryotic translation initiation
factor 4E-binding protein; PKC: protein kinase C; SGK: serum-and glucocorticoid-induced protein kinase.

insulin receptor substrate 1 (pS6K-IRS1) causing activa-
tion of AKT and survival of MM cells (Figure 1).
Interestingly, DEPTOR overexpression was confined to
myeloma cells with translocations involving cyclin D1,
D3, c-MAF, or MAFB [33]. Fernandez-Saiz et al. investi-
gated how mTORC1 and 2 interacting proteins telo-
mere maintenance 2 (Tel2) and Tel2 interacting
protein 1 (Tti1) modulate mTOR levels during starva-
tion [38]. In that study, they observed that during
energy crisis, Casein Kinase 2 (CK2) phosphorylates
Tel2 and Tti1 specifically within mTORC1 marking
them for degradation by the F-box containing protein
Fbxo9. The final result due to these events was
mTORC1 inactivation and increase in mTORC2 medi-
ated AKT phosphorylation and survival (Figure 1). The
authors observed high levels of Fbxo9 expression in a
fraction of MM patients and cell lines. Knockdown of
Fbxo9 decreased the mTORC2/AKT signaling and sur-
vival of these cells. Interestingly, Fbxo9 overexpression
was found to occur in hyperdiploid MM and not in the
other subgroups [38]. Thus, DEPTOR and Fbxo9 over-
expression appear to be mutually exclusive events and
represent two unique mechanisms through which the
PI3K/AKT/mTOR pathway gets activated in a large pro-
portion of MM patients. In addition, we suspect that
other mechanisms such as high basal levels of endo-
plasmic reticulum (ER) stress and glucose-regulated
protein 78 (GRP78) cell surface localization could also
contribute to constitutively activated AKT signaling in
MM cells [39]. Finally, we think that epigenetic
changes could also contribute to activation of this
pathway in MM cells [40].
Cross talk with other signaling pathways
In addition to the feedback loops that exist within the
PI3K/AKT/mTOR pathway, numerous other pathways
cross talk with this pathway. The most intimate and
well-documented cross talk is between the PI3K/AKT/
mTOR and RAS/RAF/MEK/ERK pathways. Depending on
the cell type and context, studies have shown that
these two pathways could cross inhibit or cross acti-
vate one another. Yu et al. showed that inhibiting ERK
with the MEK inhibitor U0126 promoted the epidermal
growth factor (EGF) stimulated association of Grb2
associated binder-1 (Gab1) with PI3K thereby activat-
ing the PI3K/AKT pathway [41]. Similarly, AKT activa-
tion phosphorylated RAF in the amino terminal
regulatory domain, which then allows 14-3-3 protein
to bind RAF and inhibit its functions [42,43]. To the

contrary, other studies have shown that both path-
ways could cross activate each other through several
mechanisms. RAS can directly interact with and acti-
vate PI3K [44,45]. In addition, RAS/RAF/MEK/ERK path-
way can feed into the PI3K/AKT/mTOR pathway at the
mTOR level [46,47]. Moreover, PI3K has been shown to
activate RAF through a Rac/PAK-dependent mechan-
ism [48]. Taken together, it is clear that these two
pathways are in a complex partnership, the outcomes
of which can vary greatly based on cellular physiology,
stress and exposure to treatment. Steinbrunn et al.
[49] observed that RAS silencing did not modulate the
PI3K/AKT/mTOR pathway in MM suggesting a lack of a
direct link between these two pathways. However,
results from preclinical studies suggest that pathway
cross inhibition could be more common in MM since
inhibiting one of the pathways with a small molecule
inhibitor leads to cross activation of the other pathway
[50,51]. Clearly, more studies are needed to (1) under-
stand if these two pathways cross talk and (2) if the
cross talk is mainly cross activating or inhibiting. In
addition to the interaction with the RAS/RAF/MEK/ERK
pathway, the PI3K/AKT/mTOR pathway cross talks with
other oncogenic pathways as well. It was reported
that AKT inhibition prevents the inhibitory phosphoryl-
ation of FOXO to up regulate receptor tyrosine kinases
such as insulin-like growth factor receptor (IGF1R),
insulin receptor (IR) and receptor tyrosine-protein kin-
ase erbB-3 (HER3) [52]. In another study, it was
observed that PI3K/AKT inhibition led to activation of
Jak2/Stat5 cascade leading to resistance [53]. The
results from our studies have suggested cross talk
between the PI3K/AKT/mTOR and Notch as well as
with Jak2/Stat3 pathways [54,55].
Preclinical studies using inhibitors of the PI3K/
AKT/mTOR pathway in MM
Much of the initial preclinical studies were performed
using the mTORC1 inhibitor rapamycin and other rapa-
logs [56]. Results from such studies showed that these
agents were able to induce a cytostatic but not a cyto-
toxic response. Even though these results were disap-
pointing, it is now clear that the lack of activity could
be attributed to several factors. First, several feedback
loops exist like those that we reviewed above that
contribute towards resistance to mTORC1 inhibitors
[57]. Second, rapamycin and rapalogs do not cause
complete inhibition of mTORC1 activity. For instance,
rapalogs are able to completely abrogate the phos-
phorylation of ribosomal protein S6 kinase (p70S6K)
and phospho ribosomal protein S6 (pS6). However,
they do not fully inhibit the phosphorylation of
AKT PATHWAY IN MYELOMA 2527
eukaryotic translation initiating factor 4E-binding pro-
tein (4EBP1) [20]. Finally, in a disease like MM where
the cells have to deal with high basal levels of ER
stress, blocking translation by mTORC1 inhibition could
promote MM cell survival by partially relieving the
high basal ER stress. Nevertheless, mTORC1 inhibition
still has promise in MM in a combination setting with
agents that promote cell death and those that could
block resistance mechanisms to mTORC1 inhibition.
mTORC1 inhibitors have shown synergy when used in
combination with existing agents such as dexametha-
sone, revlimid, and panobinostat and also other novel
agents such as the receptor tyrosine kinase inhibitor
sorafenib [58–62], heat shock protein 90 (HSP90)
inhibitor 17-AAG [63], an IGF1R inhibitor NVP-AEW541
[64], and AKT inhibitor MK2206 [50] (Table 1).
To circumvent some of the resistance mechanisms
triggered in response to rapalogs, PI3K/mTOR inhibi-
tors and mTOR kinase inhibitors have been investi-
gated in MM and these agents hold more promise as
anti-MM agents. The PI3K/mTOR inhibitor BEZ235
induced potent apoptosis in several MM cell lines as a
single agent and also enhanced activity of existing
agents such as bortezomib, doxorubicin and melpha-
lan [65,66]. Furthermore, BEZ235 was found to pro-
mote osteogenic differentiation in bone marrow
stromal cells and hence could help in alleviating bone
complications in MM [67]. However, it must be noted
that mTORC1 inhibition relieves the inhibitory phos-
phorylation of IRS1 and 2 leading to PI3K activation
and AKT phosphorylation, which may lead to resist-
ance to PI3K/mTOR inhibitors. Given the role of
mTORC2 in AKT Ser 473 phosphorylation and survival
of MM cells, mTOR kinase (mTORC1 and 2) inhibitors
have also been examined in preclinical studies. In the
first study using pp242, a mTOR kinase inhibitor,
Hoang et al. [68] observed that pp242 could inhibit
AKT phosphorylation at Ser 473 and induced more
potent proliferation inhibition and apoptosis than
rapamycin. Moreover, they also observed synergistic
cytotoxicity when combined with bortezomib. It must
be noted that rapamycin was antagonistic when used
with bortezomib [68]. We think that pAKT inhibition by
mTOR kinase inhibitors but not by mTORC1 inhibitors
could be the main factor why mTOR kinase inhibitors
but not mTORC1 inhibitors synergized with bortezomib.
A few other mTOR kinase inhibitors have been investi-
gated and results from them consistently showed
pronounced apoptosis induction in MM cells [69,70].
In spite of such promising effects, one must remember
that the feedback loop from mTORC1 to IGF1R still
exists, which could get activated post mTOR kinase
inhibitor treatment. This was noted in a study using
2528 V. RAMAKRISHNAN AND S. KUMAR

Table 1. Results from preclinical studies examining PI3K/AKT/mTOR pathway inhibitors in MM.
Drug Target Response Reference
Rapamycin and rapalogs Rapamycin: mTORC1 Cytostatic response [56]
Rapamycin plus Rapamycin: mTORC1 Synergistic cytotoxicity was observed. The combination [58]
lenalidomide Lenalidomide: retained activity in the presence of cytokines IL6, IGF-1
Immunomodulatory and bone marrow stromal cells
agent
Rapamycin plus Rapamycin: mTORC1 Synergistic apoptosis mediated by cap-dependent transla- [59]
dexamethasone Dexamethasone: tion inhibition. mTOR inhibition led to decreased transla-
corticosteroid tion of XIAP, cIAP1, HSP27, BAG3
RAD001 plus LBH589 RAD001:mTORC1 Synergistic cytotoxicity and proliferation inhibition was [60]
LBH589: Pan-Histone observed in cell lines and patient cells. Drug combin-
deacetylase (HDAC) ation caused G0/G1 arrest and inhibition of
inhibitor angiogenesis.
Rapamycin plus Rapamycin: mTORC1 Enhanced apoptosis was observed in cell lines and patient [61]
entinostat Entinostat: Class I cells. Entinostat caused up regulation of p21 and p16
HDAC inhibitor and the drug combination decreased cyclin D, Bcl-XL,
MAPK, survivin
Rapamycin plus 17- Rapamycin: mTORC1 Drug combination induced pronounced apoptosis, cell cycle [63]
N-allylamino-17-deme- inhibitor arrest, inhibited angiogenesis and osteoclast formation
thoxygeldanamycin 17-AAG: HSP90
(17-AAG) inhibitor
BEZ235 BEZ235: PI3K/mTORC1 BEZ235 induced both cell cycle arrest and cell death in [65–67]
inhibitor many myeloma cell lines and patient cells. Components
of the tumor microenvironment were unable to prevent
BEZ235 induced cell death. BEZ235 induced synergistic
cell death in combination with bortezomib, doxorubicin,
dexamethasone and melphalan. BEZ235 promoted osteo-
genic differentiation mediated by mTOR inhibition
pp242 mTORC1/2 pp242 induced potent apoptosis mediated by both rapamy- [68]
cin sensitive and insensitive target inhibition. pp242
induced synergistic apoptosis when combined with
bortezomib
INK128 mTORC1/2 INK128 induced more potent apoptosis than mTORC1 [69]
inhibitor rapamycin. Inhibition of p-4EBP1 and pAKT
were observed to be reliable biomarkers for mTORC2
inhibition.
AZD8055 mTORC1/2 AZD8055 treatment caused apoptosis in MM cells. The [70]
authors observed up regulation of IGF1R phosphorylation
after AZD8055 treatment and blocking IGF1R using an
IGF1R neutralizing antibody enhanced cell kill induced
by AZD8055
MK2206 AKT MK2206 induced potent apoptosis in some but not all cell [50]
lines. High baseline pAKT levels correlated with sensitiv-
ity to MK2206. pERK mediated activation of PI3K/AKT/
mTOR pathway downstream of AKT was observed to be
a factor contributing to resistance to MK2206. MK2206
induced synergistic cytotoxicity when used in combin-
ation with rapamycin, LY294002, MEK1/2 inhibitor and
dexamethasone
TAS-117 AKT High baseline pAKT levels correlated with sensitivity to TAS- [51]
117. TAS-117 further increased ER stress induced by pro-
teasome inhibitors leading to irrecoverable ER stress and
apoptosis
PI-103 PI-103: PI3K/mTOR Autophagy inhibition by bafilomycin A1 enhanced apop- [72]
Bafilomycin A1: tosis induced by PI-103
Autophagy inhibitor
BEZ235 BEZ235: PI3K/mTOR Chloroquine promoted BEZ235 induced apoptosis in MM [72]
Chloroquine: cells
Autophagy inhibitor

the mTOR kinase inhibitor AZD8055, where the drug
treatment led to up regulation of IGF1R and subse-
quently AKT phosphorylation at Ser 473. The authors
showed that IGF1R blocking antibody was able to
reverse the IGF1R induced pAKT and enhance cell
death induced by AZD8055 [70] (Table 1).
AKT inhibitors have also been evaluated in MM.
Perifosine, an alkylphospholipid that binds to cell
membranes and inhibits AKT, was examined in preclin-
ical studies. Perifosine induced cytotoxicity in MM cell
lines and patient cells and synergized with bortezo-
mib, dexamethasone, doxorubicin, melphalan, and
MEK1/2 inhibitor U0126 [71]. We examined the anti-
MM effects of an AKT-specific allosteric inhibitor
MK2206 and observed that cell lines with high basal
levels of pAKT were significantly more sensitive to

MK2206 [50]. Moreover, we found that cells with no to
low basal levels of pAKT still expressed similar levels of
pmTOR like those cells with high basal levels of pAKT.
We identified that constitutively active pERK in MM
cells contributed to activation of mTOR [50]. Using
MK2206 in combination with the MEK inhibitor U0126
induced synergistic cell death. In addition, MK2206
synergized with the PI3K inhibitor LY294002, rapamy-
cin, and dexamethasone in inducing cell death in MM
cells [50]. More recently, Mimura et al. [51] also
observed that the allosteric AKT inhibitor TAS-117
induced more potent cell death in cells expressing
high basal levels of pAKT. Thus, pAKT levels could
serve as a biomarker for identifying patients who are
likely to respond to AKT inhibitors. In addition, they
found that TAS-117 enhanced ER stress induced by
bortezomib and carfilzomib to induce irreversible ER
stress and enhanced apoptosis in MM cells [51]
(Table 1). mTORC1 is the most well-known inhibitor of
autophagy [20]. Targeted inhibition of mTORC1 could
therefore result in autophagy induction, and cancer
cell survival or death. In a disease like MM with abnor-
mally high levels of misfolded proteins, autophagy
might be an important mechanism through which the
cancer cells eliminate these proteins and maintain cel-
lular homeostasis. mTORC1 inhibition by either PI3K,
AKT, or mTOR inhibitors could, therefore, promote cell
survival through induction of autophagy. Aronson
et al. [72] observed autophagy induction post treat-
ment with a dual PI3K/mTOR inhibitor PI-103. By com-
bining PI-103 with an autophagy inhibitor bafilomycin
A1, the authors showed enhanced apoptosis in MM
cells. The authors observed similar effects when they
used the PI3K/mTOR inhibitor BEZ235 in combination
with a different autophagy inhibitor chloroquine [72].
In another study, it was observed that the HSP90
inhibitor 17-dimethylaminoethylamino-17-demethoxy-
geldanamycin-induced apoptosis (17-DMAG) promoted
autophagy by inhibiting mTORC1 [73]. The authors
observed enhanced apoptosis when they combined
17-DMAG with the autophagy inhibitor 3-methylade-
nine (3-MA) [73]. Examining AKT or mTOR inhibitors in
combination with autophagy inhibitors could, there-
fore, demonstrate more promising results in MM
patients. Although this combinatorial approach looks
promising in in vitro studies, the nutrient deprived
conditions that the tumors are exposed to in vivo
could dramatically alter the responses observed. In a
recently published study, it was shown that tumors
grown in medium deprived of all amino acids except
glutamine inhibit autophagy by up regulating
mTORC1 [74]. In this context, glutaminolysis activated
mTORC1, which inhibited autophagy and promoted
AKT PATHWAY IN MYELOMA 2529
apoptosis of tumor cells [74]. It remains to be seen if
mTORC1 could similarly function as a tumor suppres-
sor in MM in nutrient deprived conditions.
Clinical trials
The results obtained from preclinical studies using inhib-
itors of the PI3K/AKT/mTOR pathway strongly supported
clinical evaluation of such agents in MM patients.
mTORC1 inhibitors were the first to be examined.
In a phase II trial, Farag et al. examined the clinical
efficacy of temsirolimus (CCI-779) in relapsed refractory
MM [75] (Table 2). Sixteen patients were enrolled in
this study and received at least two cycles of treat-
ment. The response to temsirolimus was minimal with
only one patient achieving a partial response (PR) and
five achieving minimal responses (MR). The disease
remained stable in six patients and progressed in the
remaining four patients. The time to progression (TTP)
was observed to be 4 months. The authors performed
western blots on peripheral blood mononuclear cells
(PBMCs) to check for target inhibition post treatment.
They observed correlation between p-p70S6K and
p-4EBP1 inhibition in PBMCs and response to temsiroli-
mus with target inhibition observed only in patients
who showed a MR or PR to the drug [75]. In another
study, everolimus (RAD001) was examined in a phase I
trial and the result was very similar to the study by
Farag et al. [76,77] (Table 2). However, the investiga-
tors observed down regulation of pmTOR in plasma
cells of all patients irrespective of the lack or presence
of a response [77]. These two trials showed that
mTORC1 inhibitors are reasonably well tolerated but
largely ineffective in MM. In subsequent trials, mTOR
inhibitors were examined in combination with either
bortezomib or lenalidomide. Yee et al. evaluated ever-
olimus in combination with lenalidomide in a phase I
trial on relapsed MM patients [78] (Table 2). Out of the
26 patients evaluated, 23 were considered evaluable
for response and one patient showed a complete
response (CR), four patients showed PR and 10
achieved a MR for a response rate of 65%. The drug
combination was considered to be reasonably safe.
The common grade 3 or 4 adverse events observed
were found to be thrombocytopenia (35%) and neu-
tropenia (42%). The authors performed correlative
studies using plasma cells from patients before and
after treatment. The results showed down regulation
of p-p70S6K and in addition gene expression profiling
showed that the responders expressed higher levels of
pmTOR pathway members than the non-responders
[78]. Temsirolimus in combination with bortezomib
was also evaluated in relapsed refractory MM in a
2530 V. RAMAKRISHNAN AND S. KUMAR

Table 2. Results from clinical trials using PI3K/AKT/mTOR pathway inhibitors in MM.
Phase of # of
Drug Drug target trial patients Outcomes Toxicities Reference
Temsirolimus mTORC1 Phase II 16 Partial response: 1 patient Fatigue, neutropenia, [75]
(CCI-779) Minimal response: 5 patient thrombocytopenia,
Stable disease: 6 patients anemia and stomatitis
Progressive disease: 4 patients
Time to progression: 4 months
Everolimus mTORC1 Phase I 17 Partial response: 1 patient Thrombocytopenia, [76,77]
Minimal response: 1 patient leukopenia, anemia
Stable disease: 8 patients
Everolimus þ mTORC1 þ Phase I 23 Complete response: 1 patient Thrombocytopenia and [78]
lenalidomide Immunomodulator Partial response: 4 patients neutropenia. The drug
Minimal response: 10 patients combination was con-
sidered to be reason-
ably safe.
Temsirolimus þ Phase I 20 Very good partial response: 1 patient Thrombocytopenia, lym- [79]
Bortezomib Partial response: 1 patient phopenia, neutro-
Minimal response: 2 patients penia, leucopenia,
Stable disease: 12 patients anemia and diarrhea
Progressive disease: 1 patient
Temsirolimus þ Borte- Phase II 43 Complete response: 2 patients Thrombocytopenia, lym- [79]
zomib Very good partial response: 4 phopenia, neutro-
patients penia, leucopenia,
Partial response: 8 patients anemia and diarrhea
Minimal response: 6 patients
Stable disease: 19 patients
Progressive disease: 1 patient
TAK-228 mTORC1/2 Phase I 31 Minimal response: 1 patient Thrombocytopenia, [80]
Stable disease: 14 patients fatigue, neutropenia,
Progressive disease: 11 patients anemia, nausea,
hyperglycemia
Perifosine þ AKT Phase I/II 73 Complete response: 3 patients Thrombocytopenia, neu- [81]
Bortezomib ± Partial response: 13 patients tropenia, anemia and
Dexamethasone Minimal response: 14 patients pneumonia
Stable disease: 30 patients
Perifosine þ Phase I 30 Near complete response: 4 patients Thrombocytopenia, [82]
Lenalidomide þ Very good partial response: 3 leucopenia, lympho-
Dexamethasone patients penia, hyperglycemia,
Partial response: 8 patients hypophosphatemia,
Minimal response: 7 patients arthralgia, fatigue,
Stable disease: 6 patients diarrhea, nausea
Progressive disease: 2 patients
Perifosine Phase III Discontinued due to non-AKT related
toxicities
Afuresitib AKT 34 Partial response: 3 patients Neutropenia, rash, ody- [83]
Minimal response: 3 patients nophagia, fatigue,
Stable disease: 14 patients thrombocytopenia,
nausea, diarrhea
Afuresitib þ AKT, MEK/ERK Not tolerated [83,84]
Trametinib

phase 1/2 trial [79] (Table 2). In this study, it was
observed that one-third of the patients achieved at
the very least a PR. Grade 3–4 adverse events
observed were thrombocytopenia, lymphopenia, neu-
tropenia, leucopenia, anemia, and diarrhea [79]. Given
the promising preclinical studies seen using mTORC1/2
inhibitors, Ghobrial et al. [80] investigated the effects
of TAK-228, a mTORC1/2 inhibitor in relapsed refrac-
tory MM in a phase I trial (Table 2). The results
obtained were disappointing as the response to
TAK-228 was similar to what was observed with single
agent temsirolimus or everolimus. It remains to be
seen if TAK-228 or other mTORC1/2 inhibitors demon-
strate better activity when used in combination with
other existing therapies. The best response yet with
PI3K/AKT/mTOR pathway inhibitors has been with AKT
inhibitors [81,82]. Perifosine was examined in heavily
pretreated relapsed refractory MM patients in combin-
ation with bortezomib with or without dexamethasone
in a phase I/II trial [81]. The overall response rate
(ORR) defined as MR or better was 41% with 4% CR,
18% PR. The duration of response was over 6 months
for all patients showing a response. Patients who pro-
gressed on treatment or those with stable disease
were given dexamethasone and the ORR of those
patients was 38% (2%CR, 6PR, and 10MR). The most
common grade 3 or toxicities included thrombocyto-
penia, neutropenia, anemia, and pneumonia. In
another study, perifosine was investigated in a phase I
trial in combination with lenalidomide and dexametha-
sone in relapsed refractory MM patients [82]. The
results obtained were very promising with an ORR of

73% and 50% of them with a PR or better. The median
progression-free survival was 10.8 months and median
overall survival was 30.6 months. Furthermore, the
authors observed that response positively correlated
with increased levels of pAKT suggesting that pAKT
could serve as a biomarker in predicting responses
[82]. Given the very promising results with perifosine,
a large phase III trial was initiated (NCT01002248).
However, the trial was discontinued due to non-AKT-
related toxicities associated with perifosine treatment.
Despite the huge disappointment, perifosine demon-
strated the potential that AKT inhibition has in MM
treatment and how we could target the PI3K/AKT/
mTOR pathway effectively to achieve meaningful
responses in MM. However, it must be noted that a
clinical trial using the AKT-specific inhibitor afuresertib
as a single agent showed minimal response and a trial
using afuresertib in combination with the MEK inhibitor
trametinib was not well tolerated [83,84]. Even though
targeting the PI3K/AKT/mTOR and MEK/ERK pathways
looked promising in preclinical studies, it appears that
the dosing schedule could be crucial in minimizing toxic-
ities associated with such combinations [84].
Thus, despite promising pre-clinical data, inhibitors
of the PI3K/AKT/mTOR pathway have so far seen lim-
ited clinical activity in MM.
Conclusions
Basic and translational studies performed on the PI3K/
AKT/mTOR pathway in MM have shown that it plays
an integral role in MM disease biology. However, it still
remains unclear if one or multiple targets need to be
inhibited within this pathway to achieve durable
responses. The various feedback loops that exist within
this pathway and between this and the other signaling
pathways could complicate the type and depth of
response that we see in patients. In addition, the
heterogenous nature of MM makes this even more
complicated as there could be several unique mecha-
nisms that contribute to AKT pathway activation, the
type of feedback loop and the extent of cross talk
with other signaling pathways. However, with more
focused studies addressing each of these concerns and
the development of more novel and potent small mol-
ecule inhibitors of this pathway, we remain optimistic
that such drugs will help us in controlling and possibly
curing this devastating malignancy.
Potential conflict of interest: Disclosure forms provided
by the authors are available with the full text of this
article online at https://doi.org/10.1080/10428194.2017.
1421760.
AKT PATHWAY IN MYELOMA 2531
ORCID
Vijay Ramakrishnan http://orcid.org/0000-0003-3521-102X
Shaji Kumar http://orcid.org/0000-0001-5392-9284
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