Lysosomes are acidic organelles that contain hydrolytic enzymes that mediate the intracellular degradation of macromolecules. Damage of these organelles often results in lysosomal membrane permeabilization (LMP) and the release into the cytoplasm of the soluble lysosomal contents, which include proteolytic enzymes of the cathepsin family. This, in turn, activates several intracellular cascades that promote a type of regulated cell death, called lysosome-dependent cell death (LDCD). LDCD can be inhibited by pharmacological or genetic blockade of cathepsin activity, or by protecting the lysosomal membrane, thereby stabilizing the organelle. Lysosomal alterations are common in cancer cells and may increase the sensitivity of these cells to agents that promote LMP. In this review, we summarize recent findings supporting the use of LDCD as a means of killing cancer cells.

Lysosomes

The lysosome is an acidic organelle that acts as the ‘stomach’ of the cell [1]. Lysosomes contain over 60 hydrolytic enzymes that mediate the degradation of extracellular components that are delivered via endocytosis and phagocytosis or intracellular material that is delivered to the lysosome via autophagy [2]. After degradation, the resulting products are recycled back to the cytosol via transport channels or released into the extracellular space via exocytosis. While initially considered solely as a cellular garbage disposal system, recent findings have uncovered additional roles of lysosomes as essential cellular signaling platforms [3].

Lysosomal enzymes digest cellular compounds in an acidic environment (the pH of the lysosomal lumen pH is 4.5–5), but some, such as cathepsin B, D, and L, are also active at higher pH, although their stability and activity vary among them [4].

Proper localization of lysosomal enzymes within the lysosome is critical to maintain a correct cellular homeostasis, as their leakage into the cytoplasm can cause cell death by degrading essential cellular components and/or activating apoptotic pathways. This leakage is the consequence of lysosomal membrane permeabilization (LMP), a multi-pathway process that results in lysosome-dependent cell death (LDCD), which, in most cases, can be prevented by pharmacological or genetic blockade of protease activity [5,6].

LMP and LDCD

Cell death as a result of lysosomal rupture was first described by the discoverer of lysosomes, Christian de Duve, who defined them as ‘suicide bags’, owing to their content of powerful hydrolytic enzymes [7]. Although the concept was defined almost 40 years ago, this regulated cell death pathway remained poorly understood due to limitations in the methodologies used to study it. However, the recent development of new techniques has rekindled interest in this field [8,9].

LMP is a perturbation of the lysosomal membrane that allows translocation of the lysosomal contents, including lysosomal enzymes, to the cytoplasm of the cell [5,10]. Massive lysosomal rupture induces the release of the entire contents of the lysosome, triggering a cascade of hydrolysis of the cytoplasmic contents, and resulting in generalized cytoplasmic acidification with lethal consequences for the cell [6]. In contrast, partial and selective LMP triggers a cascade of regulated cell death mediated by the release of specific lysosomal enzymes into the cytosol [1114] (Figure 1). Among them, Cathepsin B and D are the main active proteases after LMP, but other lysosomal proteases such as chymotrypsin B and proteinase 3 have also been implicated in LDCD [15,16].

Inducers of LMP.

Figure 1.
Inducers of LMP.

Agents such as CADs can induce LMP and the translocation to the cytoplasm of lysosomal hydrolases (e.g. cathepsins). ROS can pass through the lysosomal membrane and, in the presence of free iron, catalyze Fenton reactions to produce highly toxic intermediates that damage lysosomal proteins, including Hsp70. Lysosomotropic detergents and some antibiotics can enter lysosomes and destabilize the lysosomal membrane, a process exacerbated by photodamage. Calpains, activated by increases in calcium, target several lysosomal proteins, including LAMP2 and Hsp70. This process is enhanced by Hsp70 oxidation caused by intralysosomal Fenton reactions. LMP is also induced by other agents, including bacterial and viral products, silica crystals, and nanoparticles.

Figure 1.
Inducers of LMP.

Agents such as CADs can induce LMP and the translocation to the cytoplasm of lysosomal hydrolases (e.g. cathepsins). ROS can pass through the lysosomal membrane and, in the presence of free iron, catalyze Fenton reactions to produce highly toxic intermediates that damage lysosomal proteins, including Hsp70. Lysosomotropic detergents and some antibiotics can enter lysosomes and destabilize the lysosomal membrane, a process exacerbated by photodamage. Calpains, activated by increases in calcium, target several lysosomal proteins, including LAMP2 and Hsp70. This process is enhanced by Hsp70 oxidation caused by intralysosomal Fenton reactions. LMP is also induced by other agents, including bacterial and viral products, silica crystals, and nanoparticles.

Pore formation in the lysosomal membrane is the simplest mean of triggering cathepsin release into the cytoplasm. Several venoms and bacterial toxins have this effect and induce apoptotic cell death [17,18]. Recent findings indicate that in addition to bacterial and viral proteins, endogenous toxin-like molecules, such as aerolysin-like proteins, are also capable of inducing LMP [19]. These molecules can translocate to lysosomes, forming high molecular-mass detergent-stable oligomers that promote lysosomal destabilization and cathepsin B release.

Lysosomotropic detergents are compounds that can cross membranes and remain trapped within the lysosome after protonation, from where they induce LMP [20]. Examples include l-leucyl-l-leucine methyl ester [21,22], the cationic amphiphilic drug (CAD) siramesine, which it has been described to detach acid sphingomyelinase (ASM) from the lysosomal membrane, promoting lysosomal destabilization [23,24] and O-methyl-serine dodecylamide hydrochloride (MSDH) [25,26], all of which induce LDCD.

LMP can also be induced in several other circumstances. For example, an increase in free-radical levels can provoke the lysosomal membrane destabilization. Lysosomes contain high concentrations of iron as a consequence of the enzymatic digestion of iron-containing protein complexes. Hydrogen peroxide, which readily diffuses through membranes, can react with intralysosomal iron in the Fenton reaction, resulting in the formation of highly reactive hydroxyl radicals that cause membrane damage, inducing LMP. In contrast, deferoxamine (DFO)-induced iron chelation can protect lysosomal membranes and reduce cell death in many conditions [2730].

The lipid composition of lysosome membranes is a key factor regulating lysosome membrane stability. Heat shock protein 70 (Hsp70) promotes the binding between the lipid bis(monoacylglycero)phosphate (BMP) and ASM. This interaction increases ASM activity and lysosomal stability [31]. Interestingly, a rise in cholesterol levels leads to the sequestration of BMP [31,32]. Interestingly, ASM mediates the degradation of sphingomyelinase to ceramide and ASM activity, or decreases in sphingomyelin levels reduce lysosomal membrane stability [33].

The activation of calpains, a group of cytosolic calcium-activated cysteine proteases, has often been associated with LMP triggering cell death under various conditions, for example in neuronal tissue after ischemia [34,35], in mouse models of retinal dystrophies [36] and also in physiological conditions as during mammary gland involution [37]. The molecular mechanisms have been associated with the cleavage of several lysosomal proteins such as Hsp70, LAMP2 that result in lysosomal membrane destabilization [34,38]. Together, these observations suggest the existence of a calcium–calpain cascade responsible for LMP and cell death [39].

LMP is not an in vitro artifact induced solely by altering lysosomal homeostasis in laboratory conditions [6]. Cell death following LMP has been observed in several physiological settings, including during mammary gland involution [37,40], during cell death associated with neutrophil regulation [16], and in bacteria-infected macrophages to prevent the spread of infection [41]. LMP has also been implicated in certain pathologies. For example, cathepsin B is released from lysosomes in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of Parkinson's disease [42,43], and calcium accumulation into photoreceptors promotes LMP and lysosomal cell death in the rd10 mouse model of retinitis pigmentosa [36]. Lysosome destabilization was also observed in a mouse model of Niemann Pick type A disease, in which neuronal cells exhibit LMP [44], although other report using fibroblasts from NPA patients did not show signs of LMP [45]. Whether LMP contributes to NPA pathology is still unknown. Given the implication of LMP in several diseases, researchers have sought to identify ways to stabilize the lysosomal membrane. For example, Hsp70 stabilizes in vivo the lysosomal membrane in Niemann Pick disease types A and B [31]. Promoting membrane stability may therefore represent a new therapeutic strategy for certain neurodegenerative conditions.

LMP can induce cell death through several pathways (Figure 2), including caspase-dependent cell death or apoptosis, as in some cases, cathepsins can activate caspases directly or in a caspase-independent manner. [5,14,46,47]. Furthermore, LMP can activate the inflammasome, inducing cytokine release and activating macrophages via a process known as pyroptosis [48]. A key characteristic of LMP-induced cell death is that in most cases can be blocked by cathepsin inhibition by using, for example, CA-074Me and E64-d to inhibit cathepsin B and pepstatin A to inhibit cathepsin D-dependent cell death, respectively [14,40].

LDCD and apoptosis are triggered after LMP.

Figure 2.
LDCD and apoptosis are triggered after LMP.

LDCD is triggered by several inducers, resulting in the translocation of cathepsins to the cytoplasm and subsequent cell death. LMP can also be induced by oxidative stress (reactive oxygen species; ROS) and can generate an amplification loop, further exacerbating both LMP and MMP. ROS and cathepsins may also regulate cell death and trigger the classical apoptotic pathway resulting in cytochrome release, caspase activation, and cell death.

Figure 2.
LDCD and apoptosis are triggered after LMP.

LDCD is triggered by several inducers, resulting in the translocation of cathepsins to the cytoplasm and subsequent cell death. LMP can also be induced by oxidative stress (reactive oxygen species; ROS) and can generate an amplification loop, further exacerbating both LMP and MMP. ROS and cathepsins may also regulate cell death and trigger the classical apoptotic pathway resulting in cytochrome release, caspase activation, and cell death.

Over the years, research has highlighted the difficulty in classifying specific cell-death subroutines based on the features of dying cells, as distinct initiation cascades can result in similar cell death morphotypes. In the case of LDCD, this issue is further complicated by the fact that lysosomal permeabilization does not produce concrete morphological alterations that can be identified by electron microscopy [49]. Some of the most powerful tools used to study LMP have only recently been developed and will probably facilitate the discovery in the near future of new conditions associated with LDCD [8]. These tools include the galectin-3 assay, which enables the identification of permeabilized lysosomes using specific antibodies, such as anti-galectin-3. Galectins are glycoprotein-binding proteins that gain access to the luminal side of the heavily glycosylated lysosomal membrane proteins after lysosomal permeabilization [8].

Lysosomal alterations in cancer

Rapidly dividing cancer cells are highly dependent on proper lysosomal function. Accordingly, transformation and cancer progression involve dramatic changes in the volume, composition, and cellular distribution of lysosomes [50]. Many of these changes provide transformed cells with selective advantages, promoting invasive growth, angiogenesis, and drug resistance. The same changes can, however, markedly sensitize cells to LMP and therefore to the effects of anticancer drugs that target the lysosome [51]. Among the changes observed in cancer cells is an increase in the levels of lysosomal enzymes. For example, the expression of heparanase, an endo-β-d-glucuronidase that participates in extracellular matrix remodeling, is increased in several cancers, including gastric carcinoma and lung and breast cancer, in which it facilitates remodeling of the extracellular matrix to promote metastasis [52,53].

Cathepsins are also frequently overexpressed in cancer cells. Increased expression of cathepsin B has been proposed as a marker of colorectal cancer in some populations [54]. The expression of cathepsin D is also increased in several tumor types, including melanoma, glioma, and lung cancer, and contributes to tumor progression in each of these scenarios [55,56].

In cancer cells, lysosomes exhibit alterations in morphology and intracellular distribution, and are more vulnerable to LMP owing to their larger size [51,57,58]. Accordingly, tumor cells may be more sensitive that non-transformed cells to the toxic effects of lysosomotropic agents [59], which are being investigated as potential anticancer agents. The pharmacological or genetic inhibition of endogenous stabilizers of the lysosomal membrane (e.g. Hsp70) is another potential means of killing cancer cells. Supporting this view, the response to LMP-based therapies is correlated with Hsp70 levels in several cancer cells lines [60]. Interestingly, expression of SMPD1, the gene encoding ASM, is decreased in gastrointestinal, hepatocellular, salivary gland, renal, and head and neck carcinomas, further sensitizing lysosomes to the effects of destabilizing agents [51].

Strategies to induce lysosomal cell death in cancer cells

As discussed above, lysosomes may constitute a powerful weapon with which to kill cancer cells from the inside. Selective induction of LMP in cancer cells could prove an efficient means of inducing cancer cell death. A better understanding of the role of lysosomal proteins and lipid composition in cancer cells could facilitate the development of more powerful drugs to kill tumors through the induction of LMP and LDCD. Below, we describe several strategies to kill cancer cells that are currently being studied.

Cationic amphiphilic drugs

Several research groups have focused their efforts on developing CADs to induce LMP and cathepsin release from cancer cells. These molecules exhibit more effects in cancer cells due to lipidomic alterations during the tumorigenic process [23,51]. CADs such as antidepressants (siramesine), anti-malarials, and antihistamines can enter lysosomes and become trapped after their protonation, leading to the accumulation in this compartment. CADs are capable of altering the lipid profile by inducing the detachment of ASM from the lysosomal membrane [61]. The increase in ASM levels within the lysosome results in cathepsin release and lysosomal cell death. The efficacy of these drugs has been already demonstrated in non-small cell lung cancer, in which antihistamine CADs such as loratadine and astemizole reduce patient mortality [62]. Terfenadine, another antihistamine CAD, induces cell death in prostate cancer cells [63], while astemizole reduces cell proliferation in breast tumor-derived cells and in leukemia cancer cells both in vitro and in vivo [63]. It is worth mentioning that CADs might be beneficial in the treatment of multi-drug-resistant cancers [62,64,65]. Taken together, these results suggest that CADs may hold significant promise as anticancer therapies.

Microtubule-destabilizing agents

Microtubules facilitate the movement of the lysosome through the cell and constitute another target through which lysosomal stability and function can be altered. Disturbances in microtubule dynamics result in lysosomal instability. For example, vincristine, a microtubule-destabilizing antimitotic drug, induces lysosomal destabilization in both HeLa and MCF7 cell lines [58]. Vincristine treatment induces cell death accompanied by cell cycle arrest and cathepsin release. Interestingly, activation of Bax and caspase and cytochrome c release, three hallmarks of apoptosis, were found in dying cells [58]. Another microtubule-destabilizer, paclitaxel, inhibits lysosomal trafficking and induces the lysosome expansion, leading to cathepsin release and LDCD in several cancer cell lines [66]. A study to identify microtubule proteins implicated in LDCD demonstrated that the depletion or pharmacological inhibition of the molecular motor KIF11/Eg5 induced LMP [59]. These proteins have now been proposed as potential drug targets, but further research needs to be done to understand the process of LMP after microtubule destabilization and whether they could be applied for cancer therapy.

Autophagy induction and cathepsin maturation

The findings of a recent study describe inhibition of cathepsin maturation as a novel means of treating cancer cells. Liu et al. [67] studied the mechanism of cell death induced by IMB-6G, an anti-inflammatory natural derivative of sophoridine, in two pancreatic cancer cell lines. Their data show that IMB-6G induced apoptosis via LMP. Interestingly, silencing of Atg5 expression attenuated the lysosomal release of cathepsin B into the cytosol and the apoptosis induced by IMB-6 treatment, suggesting that increased autophagy activity was detrimental for cell survival [67]. However, the molecular link between cathepsin maturation and lysosomal stability remains elusive and further studies are needed to clarify this point.

Endoplasmic reticulum stress-inducing compounds

Several lines of evidence indicate a link between endoplasmatic reticulum (ER) stress and lysosomal destabilization. For example, in human glioma cells, tetrahydrocannabinol (THC), the main active compound of cannabis, promotes autophagy activation, sphingolipid synthesis, and ER stress [68]. The treatment with THC induced de novo ceramide synthesis and ER stress that induces increased phosphorylation of eIF2α. Mutant of eIF2α attenuated this response. Interestingly, eIF2α increased the mRNA expression of ER stress-response genes such as ATF4, CHOP, and TRB3 [68]. Later studies demonstrated that THC treatment induced alterations in sphingolipid composition in the ER, which resulted in lysosomal destabilization, cathepsin release, and LMP [68,69]. Additionally, in their study of the relationship between ER stress and cell death in pancreatic cancer, Dauer et al. [70] found that inhibition of the transcription factor-specific protein 1, Sp1 (a key component of the unfolded protein response), induced LMP in the MiaPaCa-2 pancreatic cell line. These findings indicate that ER stress promotes lysosomal destabilization, although the underlying molecular mechanisms remain to be unraveled.

Histone deacetylases and sirtuins

The sirtuin family is a group of NAD(+)-dependent deacetylases [71]. In 2014, Lahusen and Deng [72] studied the effects of SRT1720, a SIRT1 activator, in the MDA-MB-231 breast cancer cell line, and found that SRT1720 treatment induced cell death independently of Sirt1. While SRT1720-induced cell death was preceded by autophagy activation as a protective mechanism, the authors reported that cell death was inhibited with lysosomal acidity inhibitors, suggesting the implication of lysosomes during cell death under these conditions.

Calcium-dependent LMP pathways

Lysosomes act as essential calcium stores and play an important role in calcium signaling [73,74], modulating calcium release through multiple calcium channels in the lysosomal membrane. As previously mentioned above, calpains, which are calcium-activated proteases, are capable of cleaving Hsp70, inducing lysosomal destabilization [34]. Additionally, in their study in a gastric cancer cell line, Lv et al. [75] found that mushroom-derived hispidin induced cell death via caspase 9 cleavage. This cell death could not be rescued with the general caspase inhibitor z-VAD. Further analysis of the underlying mechanism revealed that hispidin induced LMP. While cathepsins were not implicated in this cell death process, treatment with the calcium chelator BAPTA restored cell viability after hispidin treatment, indicating a role of lysosomal calcium. This result suggests that lysosomal calcium may trigger cell death after the induction of LMP. Whether this effect is associated with calcium-induced damage to other organelles such as mitochondria and ER, or mediated by calpain activation, remains to be elucidated [34,37].

Mitochondrial membrane permeabilization

In many instances, LMP occurs upstream of mitochondrial membrane permeabilization (MMP), and cells deficient in Bax and Bak are completely resistant to LDCD induced by several antibiotics and lysosomotropic agents [47,76]. Interestingly, LMP still occurred in Bax and Bak double deficient cells, demonstrating that LMP is upstream of MMP [47,76]. In the HaCaT skin cancer cell line, siramesine at high concentrations induces MMP and cell death in an LMP-independent manner [77]. Conversely, and in agreement with siramesine acting as a CAD, low doses of siramesine result in a lower rate of cell death that occurs in a lysosome-dependent manner [77,78]. In other circumstances, MMP appears to occur upstream of lysosomal damage. For example, following combined treatment with tumor necrosis factor alpha (TNF-α) and cycloheximide, activation of caspase 8 and caspase 3 is observed 1 h after treatment, while cathepsin B and cathepsin D are detected 6 h after treatment [79]. While these effects could reflect differences in the sensitivity in the detection methods for caspases and cathepsin activity, these findings suggest that LMP is induced by ROS (reactive oxygen species) generated by MMP and this can represent an amplifying loop in the cell death cascade. It is also worth mentioning that TNF-α effects are highly cell-type dependent. In future studies, it will be essential to identify the pathways and processes that lead to MMP or LMP as the first step in the induction of cell death and whether the MMP–LMP axis could be exploited for therapeutic purposes.

Lysosomal photodamage

The lethal effects of photodynamic therapy on cancer cells, including breast cancer cells [80,81], are well described [80,81]. Photodamage induces both LMP and cell death [82,83]. Interestingly, lysosomes become more resistant to photodamage with increasing intraluminal pH, suggesting that the toxic effects of photodamage are dependent on lysosomal pH [84].

Perspectives

LDCD can be induced by targeting several processes, including autophagy, cathepsin maturation, endoplasmic stress, MMP, and lysosomal calcium release. While a large variety of stimuli have been implicated in LMP, further research will be necessary to fully understand the mechanisms that underlie this process. Moreover, a greater knowledge of the pathways involved in lysosomal membrane stabilization, including the interaction between Hsp70 and BMP, could allow researchers to target this process to kill cancer cells. Autophagy inhibition, another consequence of lysosomal disturbance, constitutes another novel therapeutic approach for several types of malignances. Indeed, the anticancer effects of the autophagy inhibitor hydroxychloroquine combined with other chemotherapy agents have been studied in several recent clinical trials [85], for example, in renal carcinoma, pancreatic adenocarcinoma, or melanoma [86,87]. In conclusion, the targeting of lysosomes can be a novel therapeutic approach to kill cancer cells [8890].

Abbreviations

     
  • ASM

    acid sphingomyelinase

  •  
  • BMP

    bis(monoacylglycero)phosphate

  •  
  • CAD

    cationic amphiphilic drug

  •  
  • ER

    endoplasmatic reticulum

  •  
  • Hsp70

    Heat shock protein 70

  •  
  • LDCD

    lysosome-dependent cell death

  •  
  • LMP

    lysosomal membrane permeabilization

  •  
  • MMP

    mitochondrial membrane permeabilization

  •  
  • NSCLC

    non-small cell lung cancer

  •  
  • ROS

    reactive oxygen species

  •  
  • THC

    tetrahydrocannabinol

  •  
  • TNF-α

    tumor necrosis factor alpha

Author Contribution

A.S.-P. and P.B. wrote the manuscript.

Funding

Research in the P.B.'s laboratory is supported by grants BFU2015-65623 and BFU2015-71869-REDT from Spain's Ministerio de Economia, Industria y Competitividad (MINECO), COST Action Transautophagy (CA15138), and H2020-MSCA-ITN-2017 MSCA-ITN-ETN 765912. A.S.-P. received a FPI grant from MINECO.

Competing Interests

The Authors declare that there are no competing interests associated with the manuscript.

References

References
1
De Duve
,
C.
and
Wattiaux
,
R.
(
1966
)
Functions of lysosomes
.
Annu. Rev. Physiol.
28
,
435
492
2
Shen
,
H.M.
and
Mizushima
,
N.
(
2014
)
At the end of the autophagic road: an emerging understanding of lysosomal functions in autophagy
.
Trends Biochem. Sci.
39
,
61
71
3
Carroll
,
B.
and
Dunlop
,
E.A.
(
2017
)
The lysosome: a crucial hub for AMPK and mTORC1 signalling
.
Biochem. J.
474
,
1453
1466
4
Cirman
,
T.
,
Oresic
,
K.
,
Mazovec
,
G.D.
,
Turk
,
V.
,
Reed
,
J.C.
,
Myers
,
R.M.
et al. 
(
2004
)
Selective disruption of lysosomes in HeLa cells triggers apoptosis mediated by cleavage of Bid by multiple papain-like lysosomal cathepsins
.
J. Biol. Chem.
279
,
3578
3587
5
Boya
,
P.
and
Kroemer
,
G.
(
2008
)
Lysosomal membrane permeabilization in cell death
.
Oncogene
27
,
6434
6451
6
Serrano-Puebla
,
A.
and
Boya
,
P.
(
2016
)
Lysosomal membrane permeabilization in cell death: new evidence and implications for health and disease
.
Ann. N. Y. Acad. Sci.
1371
,
30
44
7
de Duve
,
C.
(
1983
)
Lysosomes revisited
.
Eur. J. Biochem.
137
,
391
397
8
Aits
,
S.
,
Kricker
,
J.
,
Liu
,
B.
,
Ellegaard
,
A.M.
,
Hamalisto
,
S.
,
Tvingsholm
,
S.
et al. 
(
2015
)
Sensitive detection of lysosomal membrane permeabilization by lysosomal galectin puncta assay
.
Autophagy
11
,
1408
1424
9
Aits
,
S.
,
Jäättelä
,
M.
and
Nylandsted
,
J.
(
2015
)
Methods for the quantification of lysosomal membrane permeabilization: a hallmark of lysosomal cell death
.
Methods Cell Biol.
126
,
261
285
10
Repnik
,
U.
,
Hafner Česen
,
M.
and
Turk
,
B.
(
2014
)
Lysosomal membrane permeabilization in cell death: concepts and challenges
.
Mitochondrion
19
,
49
57
11
Terman
,
A.
,
Kurz
,
T.
,
Gustafsson
,
B.
and
Brunk
,
U.T.
(
2006
)
Lysosomal labilization
.
IUBMB Life
58
,
531
539
12
Aits
,
S.
and
Jaattela
,
M.
(
2013
)
Lysosomal cell death at a glance
.
J. Cell Sci.
126
(
Pt 9
),
1905
1912
13
Boya
,
P.
(
2012
)
Lysosomal function and dysfunction: mechanism and disease
.
Antioxid. Redox Signal.
17
,
766
774
14
Gómez-Sintes
,
R.
,
Ledesma
,
M.D.
and
Boya
,
P.
(
2016
)
Lysosomal cell death mechanisms in aging
.
Ageing Res. Rev.
32
,
150
168
15
Zhao
,
K.
,
Zhao
,
X.
,
Tu
,
Y.
,
Miao
,
Q.
,
Cao
,
D.
,
Duan
,
W.
et al. 
(
2010
)
Lysosomal chymotrypsin B potentiates apoptosis via cleavage of Bid
.
Cell. Mol. Life Sci.
67
,
2665
2678
16
Loison
,
F.
,
Zhu
,
H.
,
Karatepe
,
K.
,
Kasorn
,
A.
,
Liu
,
P.
,
Ye
,
K.
et al. 
(
2014
)
Proteinase 3-dependent caspase-3 cleavage modulates neutrophil death and inflammation
.
J. Clin. Invest.
124
,
4445
4458
17
Matsuda
,
S.
,
Okada
,
N.
,
Kodama
,
T.
,
Honda
,
T.
and
Iida
,
T.
(
2012
)
A cytotoxic type III secretion effector of Vibrio parahaemolyticus targets vacuolar H+-ATPase subunit c and ruptures host cell lysosomes
.
PLoS Pathog.
8
,
e1002803
18
Bewley
,
M.A.
,
Naughton
,
M.
,
Preston
,
J.
,
Mitchell
,
A.
,
Holmes
,
A.
,
Marriott
,
H.M.
et al. 
(
2014
)
Pneumolysin activates macrophage lysosomal membrane permeabilization and executes apoptosis by distinct mechanisms without membrane pore formation
.
MBio
5
,
e01710
14
19
Xiang
,
Y.
,
Yan
,
C.
,
Guo
,
X.
,
Zhou
,
K.
,
Li
,
S.
,
Gao
,
Q.
et al. 
(
2014
)
Host-derived, pore-forming toxin-like protein and trefoil factor complex protects the host against microbial infection
.
Proc. Natl Acad. Sci. U.S.A.
111
,
6702
6707
20
Miller
,
D.K.
,
Griffiths
,
E.
,
Lenard
,
J.
and
Firestone
,
R.A.
(
1983
)
Cell killing by lysosomotropic detergents
.
J. Cell Biol.
97
,
1841
1851
21
Uchimoto
,
T.
,
Nohara
,
H.
,
Kamehara
,
R.
,
Iwamura
,
M.
,
Watanabe
,
N.
and
Kobayashi
,
Y.
(
1999
)
Mechanism of apoptosis induced by a lysosomotropic agent, l-leucyl-l-leucine methyl ester
.
Apoptosis
4
,
357
362
22
Maejima
,
I.
,
Takahashi
,
A.
,
Omori
,
H.
,
Kimura
,
T.
,
Takabatake
,
Y.
,
Saitoh
,
T.
et al. 
(
2013
)
Autophagy sequesters damaged lysosomes to control lysosomal biogenesis and kidney injury
.
EMBO J.
32
,
2336
2347
23
Petersen
,
N.H.
,
Olsen
,
O.D.
,
Groth-Pedersen
,
L.
,
Ellegaard
,
A.M.
,
Bilgin
,
M.
,
Redmer
,
S.
et al. 
(
2013
)
Transformation-associated changes in sphingolipid metabolism sensitize cells to lysosomal cell death induced by inhibitors of acid sphingomyelinase
.
Cancer Cell
24
,
379
393
24
Ostenfeld
,
M.S.
,
Hoyer-Hansen
,
M.
,
Bastholm
,
L.
,
Fehrenbacher
,
N.
,
Olsen
,
O.D.
,
Groth-Pedersen
,
L.
et al. 
(
2008
)
Anti-cancer agent siramesine is a lysosomotropic detergent that induces cytoprotective autophagosome accumulation
.
Autophagy
4
,
487
499
25
Wilson
,
P.D.
,
Firestone
,
R.A.
and
Lenard
,
J.
(
1987
)
The role of lysosomal enzymes in killing of mammalian cells by the lysosomotropic detergent N-dodecylimidazole
.
J. Cell Biol.
104
,
1223
1229
26
Li
,
W.
,
Yuan
,
X.
,
Nordgren
,
G.
,
Dalen
,
H.
,
Dubowchik
,
G.M.
,
Firestone
,
R.A.
et al. 
(
2000
)
Induction of cell death by the lysosomotropic detergent MSDH
.
FEBS Lett.
470
,
35
39
27
Kurz
,
T.
,
Gustafsson
,
B.
and
Brunk
,
U.T.
(
2006
)
Intralysosomal iron chelation protects against oxidative stress-induced cellular damage
.
FEBS J.
273
,
3106
3117
28
Castino
,
R.
,
Fiorentino
,
I.
,
Cagnin
,
M.
,
Giovia
,
A.
and
Isidoro
,
C.
(
2011
)
Chelation of lysosomal iron protects dopaminergic SH-SY5Y neuroblastoma cells from hydrogen peroxide toxicity by precluding autophagy and Akt dephosphorylation
.
Toxicol. Sci.
123
,
523
541
29
Krenn
,
M.A.
,
Schürz
,
M.
,
Teufl
,
B.
,
Uchida
,
K.
,
Eckl
,
P.M.
and
Bresgen
,
N.
(
2015
)
Ferritin-stimulated lipid peroxidation, lysosomal leak, and macroautophagy promote lysosomal “metastability” in primary hepatocytes determining in vitro cell survival
.
Free Radic. Biol. Med.
80
,
48
58
30
Terman
,
A.
and
Kurz
,
T.
(
2013
)
Lysosomal iron, iron chelation, and cell death
.
Antioxid. Redox Signal.
18
,
888
898
31
Kirkegaard
,
T.
,
Roth
,
A.G.
,
Petersen
,
N.H.
,
Mahalka
,
A.K.
,
Olsen
,
O.D.
,
Moilanen
,
I.
et al. 
(
2010
)
Hsp70 stabilizes lysosomes and reverts Niemann–Pick disease-associated lysosomal pathology
.
Nature
463
,
549
553
32
Toops
,
K.A.
,
Tan
,
L.X.
,
Jiang
,
Z.
,
Radu
,
R.A.
and
Lakkaraju
,
A.
(
2015
)
Cholesterol-mediated activation of acid sphingomyelinase disrupts autophagy in the retinal pigment epithelium
.
Mol. Biol. Cell
26
,
1
14
33
Petersen
,
N.H.
,
Kirkegaard
,
T.
,
Olsen
,
O.D.
and
Jäättelä
,
M.
(
2010
)
Connecting Hsp70, sphingolipid metabolism and lysosomal stability
.
Cell Cycle
9
,
2305
2309
34
Sahara
,
S.
and
Yamashima
,
T.
(
2010
)
Calpain-mediated Hsp70.1 cleavage in hippocampal CA1 neuronal death
.
Biochem. Biophys. Res. Commun.
393
,
806
811
35
Yamashima
,
T.
and
Oikawa
,
S.
(
2009
)
The role of lysosomal rupture in neuronal death
.
Prog. Neurobiol.
89
,
343
358
36
Rodriguez-Muela
,
N.
,
Hernandez-Pinto
,
A.M.
,
Serrano-Puebla
,
A.
,
Garcia-Ledo
,
L.
,
Latorre
,
S.H.
,
de la Rosa
,
E.J.
et al. 
(
2015
)
Lysosomal membrane permeabilization and autophagy blockade contribute to photoreceptor cell death in a mouse model of retinitis pigmentosa
.
Cell Death Differ.
22
,
476
487
37
Arnandis
,
T.
,
Ferrer-Vicens
,
I.
,
Garcia-Trevijano
,
E.R.
,
Miralles
,
V.J.
,
Garcia
,
C.
,
Torres
,
L.
et al. 
(
2012
)
Calpains mediate epithelial-cell death during mammary gland involution: mitochondria and lysosomal destabilization
.
Cell Death Differ.
19
,
1536
1548
38
Villalpando Rodriguez
,
G.E.
and
Torriglia
,
A.
(
2013
)
Calpain 1 induce lysosomal permeabilization by cleavage of lysosomal associated membrane protein 2
.
Biochim. Biophys. Acta
1833
,
2244
2253
39
Yamashima
,
T.
(
2013
)
Reconsider Alzheimer's disease by the ‘calpain-cathepsin hypothesis’—a perspective review
.
Prog. Neurobiol.
105
,
1
23
40
Kreuzaler
,
P.A.
,
Staniszewska
,
A.D.
,
Li
,
W.
,
Omidvar
,
N.
,
Kedjouar
,
B.
,
Turkson
,
J.
et al. 
(
2011
)
Stat3 controls lysosomal-mediated cell death in vivo
.
Nat. Cell Biol.
13
,
303
309
41
Zhu
,
W.
,
Tao
,
L.
,
Quick
,
M.L.
,
Joyce
,
J.A.
,
Qu
,
J.M.
and
Luo
,
Z.Q.
(
2015
)
Sensing cytosolic RpsL by macrophages induces lysosomal cell death and termination of bacterial infection
.
PLoS Pathog.
11
,
e1004704
42
Dehay
,
B.
,
Bove
,
J.
,
Rodriguez-Muela
,
N.
,
Perier
,
C.
,
Recasens
,
A.
,
Boya
,
P.
et al. 
(
2010
)
Pathogenic lysosomal depletion in Parkinson's disease
.
J. Neurosci.
30
,
12535
12544
43
Vila
,
M.
,
Bove
,
J.
,
Dehay
,
B.
,
Rodriguez-Muela
,
N.
and
Boya
,
P.
(
2011
)
Lysosomal membrane permeabilization in Parkinson disease
.
Autophagy
7
,
98
100
44
Gabandé-Rodríguez
,
E.
,
Boya
,
P.
,
Labrador
,
V.
,
Dotti
,
C.G.
and
Ledesma
,
M.D.
(
2014
)
High sphingomyelin levels induce lysosomal damage and autophagy dysfunction in Niemann Pick disease type A
.
Cell Death Differ.
21
,
864
875
45
Corcelle-Termeau
,
E.
,
Vindelov
,
S.D.
,
Hamalisto
,
S.
,
Mograbi
,
B.
,
Keldsbo
,
A.
,
Brasen
,
J.H.
et al. 
(
2016
)
Excess sphingomyelin disturbs ATG9A trafficking and autophagosome closure
.
Autophagy
12
,
833
849
46
Conus
,
S.
,
Pop
,
C.
,
Snipas
,
S.J.
,
Salvesen
,
G.S.
and
Simon
,
H.U.
(
2012
)
Cathepsin D primes caspase-8 activation by multiple intra-chain proteolysis
.
J. Biol. Chem.
287
,
21142
21151
47
Boya
,
P.
,
Andreau
,
K.
,
Poncet
,
D.
,
Zamzami
,
N.
,
Perfettini
,
J.L.
,
Metivier
,
D.
et al. 
(
2003
)
Lysosomal membrane permeabilization induces cell death in a mitochondrion-dependent fashion
.
J. Exp. Med.
197
,
1323
1334
48
Heid
,
M.E.
,
Keyel
,
P.A.
,
Kamga
,
C.
,
Shiva
,
S.
,
Watkins
,
S.C.
and
Salter
,
R.D.
(
2013
)
Mitochondrial reactive oxygen species induces NLRP3-dependent lysosomal damage and inflammasome activation
.
J. Immunol.
191
,
5230
5238
49
Brunk
,
U.T.
and
Dricsson
,
J.L.
(
1972
)
Cytochemical evidence for the leakage of acid phosphatase through ultrastructurally intact lysosomal membranes
.
Histochem. J.
4
,
479
491
50
Glunde
,
K.
,
Guggino
,
S.E.
,
Solaiyappan
,
M.
,
Pathak
,
A.P.
,
Ichikawa
,
Y.
and
Bhujwalla
,
Z.M.
(
2003
)
Extracellular acidification alters lysosomal trafficking in human breast cancer cells
.
Neoplasia
5
,
533
545
51
Kallunki
,
T.
,
Olsen
,
O.D.
and
Jäättelä
,
M.
(
2013
)
Cancer-associated lysosomal changes: friends or foes?
Oncogene
32
,
1995
2004
52
Tang
,
W.
,
Nakamura
,
Y.
,
Tsujimoto
,
M.
,
Sato
,
M.
,
Wang
,
X.
,
Kurozumi
,
K.
et al. 
(
2002
)
Heparanase: a key enzyme in invasion and metastasis of gastric carcinoma
.
Mod. Pathol.
15
,
593
598
53
Sun
,
X.
,
Zhang
,
G.
,
Nian
,
J.
,
Yu
,
M.
,
Chen
,
S.
,
Zhang
,
Y.
et al. 
(
2017
)
Elevated heparanase expression is associated with poor prognosis in breast cancer: a study based on systematic review and TCGA data
.
Oncotarget
8
,
43521
43535
54
Abdulla
,
M.H.
,
Valli-Mohammed
,
M.A.
,
Al-Khayal
,
K.
,
Al Shkieh
,
A.
,
Zubaidi
,
A.
,
Ahmad
,
R.
et al. 
(
2017
)
Cathepsin B expression in colorectal cancer in a Middle East population: Potential value as a tumor biomarker for late disease stages
.
Oncol. Rep.
37
,
3175
3180
55
Fukuda
,
M.E.
,
Iwadate
,
Y.
,
Machida
,
T.
,
Hiwasa
,
T.
,
Nimura
,
Y.
,
Nagai
,
Y.
et al. 
(
2005
)
Cathepsin D is a potential serum marker for poor prognosis in glioma patients
.
Cancer Res.
65
,
5190
5194
56
Vetvicka
,
V.
,
Vetvickova
,
J.
and
Benes
,
P.
(
2004
)
Role of enzymatically inactive procathepsin D in lung cancer
.
Anticancer Res.
24
,
2739
2743
PMID:
[PubMed]
57
Ono
,
K.
,
Kim
,
S.O.
and
Han
,
J.
(
2003
)
Susceptibility of lysosomes to rupture is a determinant for plasma membrane disruption in tumor necrosis factor alpha-induced cell death
.
Mol. Cell. Biol.
23
,
665
676
58
Groth-Pedersen
,
L.
,
Ostenfeld
,
M.S.
,
Hoyer-Hansen
,
M.
,
Nylandsted
,
J.
and
Jaattela
,
M.
(
2007
)
Vincristine induces dramatic lysosomal changes and sensitizes cancer cells to lysosome-destabilizing siramesine
.
Cancer Res.
67
,
2217
2225
59
Groth-Pedersen
,
L.
,
Aits
,
S.
,
Corcelle-Termeau
,
E.
,
Petersen
,
N.H.
,
Nylandsted
,
J.
and
Jäättelä
,
M.
(
2012
)
Identification of cytoskeleton-associated proteins essential for lysosomal stability and survival of human cancer cells
.
PLoS ONE
7
,
e45381
60
Mena
,
S.
,
Rodriguez
,
M.L.
,
Ponsoda
,
X.
,
Estrela
,
J.M.
,
Jaattela
,
M.
and
Ortega
,
A.L.
(
2012
)
Pterostilbene-induced tumor cytotoxicity: a lysosomal membrane permeabilization-dependent mechanism
.
PLoS ONE
7
,
e44524
61
Kornhuber
,
J.
,
Tripal
,
P.
,
Reichel
,
M.
,
Muhle
,
C.
,
Rhein
,
C.
,
Muehlbacher
,
M.
et al. 
(
2010
)
Functional Inhibitors of acid sphingomyelinase (FIASMAs): a novel pharmacological group of drugs with broad clinical applications
.
Cell. Physiol. Biochem.
26
,
9
20
62
Ellegaard
,
A.M.
,
Dehlendorff
,
C.
,
Vind
,
A.C.
,
Anand
,
A.
,
Cederkvist
,
L.
,
Petersen
,
N.H.
et al. 
(
2016
)
Repurposing cationic amphiphilic antihistamines for cancer treatment
.
EBioMedicine
9
,
130
139
63
Wang
,
W.T.
,
Chen
,
Y.H.
,
Hsu
,
J.L.
,
Leu
,
W.J.
,
Yu
,
C.C.
,
Chan
,
S.H.
et al. 
(
2014
)
Terfenadine induces anti-proliferative and apoptotic activities in human hormone-refractory prostate cancer through histamine receptor-independent Mcl-1 cleavage and Bak up-regulation
.
Naunyn Schmiedebergs Arch. Pharmacol.
387
,
33
45
64
Jaffrezou
,
J.P.
,
Chen
,
G.
,
Duran
,
G.E.
,
Muller
,
C.
,
Bordier
,
C.
,
Laurent
,
G.
et al. 
(
1995
)
Inhibition of lysosomal acid sphingomyelinase by agents which reverse multidrug resistance
.
Biochim. Biophys. Acta
1266
,
1
8
65
Hait
,
W.N.
,
Gesmonde
,
J.F.
,
Murren
,
J.R.
,
Yang
,
J.M.
,
Chen
,
H.X.
and
Reiss
,
M.
(
1993
)
Terfenadine (Seldane®): a new drug for restoring sensitivity to multidrug resistant cancer cells
.
Biochem. Pharmacol.
45
,
401
406
66
Castino
,
R.
,
Peracchio
,
C.
,
Salini
,
A.
,
Nicotra
,
G.
,
Trincheri
,
N.F.
,
Demoz
,
M.
et al. 
(
2009
)
Chemotherapy drug response in ovarian cancer cells strictly depends on a cathepsin D-Bax activation loop
.
J. Cell. Mol. Med.
13
,
1096
1109
67
Liu
,
L.
,
Zhang
,
N.
,
Dou
,
Y.
,
Mao
,
G.
,
Bi
,
C.
,
Pang
,
W.
et al. 
(
2017
)
Lysosomal dysfunction and autophagy blockade contribute to IMB-6G-induced apoptosis in pancreatic cancer cells
.
Sci. Rep.
7
,
41862
68
Salazar
,
M.
,
Carracedo
,
A.
,
Salanueva
,
I.J.
,
Hernandez-Tiedra
,
S.
,
Lorente
,
M.
,
Egia
,
A.
et al. 
(
2009
)
Cannabinoid action induces autophagy-mediated cell death through stimulation of ER stress in human glioma cells
.
J. Clin. Invest.
119
,
1359
1372
69
Salazar
,
M.
,
Carracedo
,
A.
,
Salanueva
,
I.J.
,
Hernandez-Tiedra
,
S.
,
Egia
,
A.
,
Lorente
,
M.
et al. 
(
2009
)
TRB3 links ER stress to autophagy in cannabinoid anti-tumoral action
.
Autophagy
5
,
1048
1049
70
Dauer
,
P.
,
Gupta
,
V.K.
,
McGinn
,
O.
,
Nomura
,
A.
,
Sharma
,
N.S.
,
Arora
,
N.
et al. 
(
2017
)
Inhibition of Sp1 prevents ER homeostasis and causes cell death by lysosomal membrane permeabilization in pancreatic cancer
.
Sci. Rep.
7
,
1564
71
Chang
,
H.C.
and
Guarente
,
L.
(
2014
)
SIRT1 and other sirtuins in metabolism
.
Trends Endocrinol. Metab.
25
,
138
145
72
Lahusen
,
T.J.
and
Deng
,
C.X.
(
2015
)
SRT1720 induces lysosomal-dependent cell death of breast cancer cells
.
Mol. Cancer Ther.
14
,
183
192
73
Raffaello
,
A.
,
Mammucari
,
C.
,
Gherardi
,
G.
and
Rizzuto
,
R.
(
2016
)
Calcium at the center of cell signaling: interplay between endoplasmic reticulum, mitochondria, and lysosomes
.
Trends Biochem. Sci.
41
,
1035
1049
74
Li
,
R.J.
,
Xu
,
J.
,
Fu
,
C.
,
Zhang
,
J.
,
Zheng
,
Y.G.
,
Jia
,
H.
et al. 
(
2016
)
Regulation of mTORC1 by lysosomal calcium and calmodulin
.
eLife
5
,
e19360
75
Lv
,
L.X.
,
Zhou
,
Z.X.
,
Zhou
,
Z.
,
Zhang
,
L.J.
,
Yan
,
R.
,
Zhao
,
Z.
et al. 
(
2017
)
Hispidin induces autophagic and necrotic death in SGC-7901 gastric cancer cells through lysosomal membrane permeabilization by inhibiting tubulin polymerization
.
Oncotarget
8
,
26992
27006
76
Boya
,
P.
,
Gonzalez-Polo
,
R.A.
,
Poncet
,
D.
,
Andreau
,
K.
,
Vieira
,
H.L.
,
Roumier
,
T.
et al. 
(
2003
)
Mitochondrial membrane permeabilization is a critical step of lysosome-initiated apoptosis induced by hydroxychloroquine
.
Oncogene
22
,
3927
3936
77
Česen
,
M.H.
,
Repnik
,
U.
,
Turk
,
V.
and
Turk
,
B.
(
2013
)
Siramesine triggers cell death through destabilisation of mitochondria, but not lysosomes
.
Cell Death Dis.
4
,
e818
78
Ostenfeld
,
M.S.
,
Fehrenbacher
,
N.
,
Hoyer-Hansen
,
M.
,
Thomsen
,
C.
,
Farkas
,
T.
and
Jaattela
,
M.
(
2005
)
Effective tumor cell death by σ-2 receptor ligand siramesine involves lysosomal leakage and oxidative stress
.
Cancer Res.
65
,
8975
8983
79
Bidovec
,
K.
,
Bozic
,
J.
,
Dolenc
,
I.
,
Turk
,
B.
,
Turk
,
V.
and
Stoka
,
V.
(
2017
)
Tumor necrosis factor-α induced apoptosis in U937 cells promotes cathepsin D-independent stefin B degradation
.
J. Cell. Biochem.
118
,
4813
4820
80
George
,
B.P.
and
Abrahamse
,
H.
(
2016
)
A review on novel breast cancer therapies: photodynamic therapy and plant derived agent induced cell death mechanisms
.
Anticancer Agents Med. Chem.
16
,
793
801
81
Li
,
H.
,
Liu
,
C.
,
Zeng
,
Y.P.
,
Hao
,
Y.H.
,
Huang
,
J.W.
,
Yang
,
Z.Y.
et al. 
(
2016
)
Nanoceria-mediated drug delivery for targeted photodynamic therapy on drug-resistant breast cancer
.
ACS Appl. Mater. Interfaces
8
,
31510
31523
82
Buytaert
,
E.
,
Dewaele
,
M.
and
Agostinis
,
P.
(
2007
)
Molecular effectors of multiple cell death pathways initiated by photodynamic therapy
.
Biochim. Biophys. Acta
1776
,
86
107
83
Caruso
,
J.A.
,
Mathieu
,
P.A.
,
Joiakim
,
A.
,
Leeson
,
B.
,
Kessel
,
D.
,
Sloane
,
B.F.
et al. 
(
2004
)
Differential susceptibilities of murine hepatoma 1c1c7 and Tao cells to the lysosomal photosensitizer NPe6: influence of aryl hydrocarbon receptor on lysosomal fragility and protease contents
.
Mol. Pharmacol.
65
,
1016
1028
84
Kessel
,
D.H.
,
Price
,
M.
and
Reiners
, Jr,
J.J.
(
2012
)
ATG7 deficiency suppresses apoptosis and cell death induced by lysosomal photodamage
.
Autophagy
8
,
1333
1341
85
Poklepovic
,
A.
and
Gewirtz
,
D.A.
(
2014
)
Outcome of early clinical trials of the combination of hydroxychloroquine with chemotherapy in cancer
.
Autophagy
10
,
1478
1480
86
Mahalingam
,
D.
,
Mita
,
M.
,
Sarantopoulos
,
J.
,
Wood
,
L.
,
Amaravadi
,
R.K.
,
Davis
,
L.E.
et al. 
(
2014
)
Combined autophagy and HDAC inhibition: a phase I safety, tolerability, pharmacokinetic, and pharmacodynamic analysis of hydroxychloroquine in combination with the HDAC inhibitor vorinostat in patients with advanced solid tumors
.
Autophagy
10
,
1403
1414
87
Wolpin
,
B.M.
,
Rubinson
,
D.A.
,
Wang
,
X.
,
Chan
,
J.A.
,
Cleary
,
J.M.
,
Enzinger
,
P.C.
et al. 
(
2014
)
Phase II and pharmacodynamic study of autophagy inhibition using hydroxychloroquine in patients with metastatic pancreatic adenocarcinoma
.
Oncologist
19
,
637
638
88
Vogl
,
D.T.
,
Stadtmauer
,
E.A.
,
Tan
,
K.S.
,
Heitjan
,
D.F.
,
Davis
,
L.E.
,
Pontiggia
,
L.
et al. 
(
2014
)
Combined autophagy and proteasome inhibition: a phase 1 trial of hydroxychloroquine and bortezomib in patients with relapsed/refractory myeloma
.
Autophagy
10
,
1380
1390
89
Rangwala
,
R.
,
Leone
,
R.
,
Chang
,
Y.C.
,
Fecher
,
L.A.
,
Schuchter
,
L.M.
,
Kramer
,
A.
et al. 
(
2014
)
Phase I trial of hydroxychloroquine with dose-intense temozolomide in patients with advanced solid tumors and melanoma
.
Autophagy
10
,
1369
1379
90
Rangwala
,
R.
,
Chang
,
Y.C.
,
Hu
,
J.
,
Algazy
,
K.M.
,
Evans
,
T.L.
,
Fecher
,
L.A.
et al. 
(
2014
)
Combined MTOR and autophagy inhibition: phase I trial of hydroxychloroquine and temsirolimus in patients with advanced solid tumors and melanoma
.
Autophagy
10
,
1391
1402