メタボ、肥満症はＰＡＫ遮断剤であるプロポリスや納豆などで治療可能かもしれない！

炎症現象には一般に、「ＰＡＫ」という発癌キナーゼが必須であることが米国で
最近明らかになった。従って、「ＣＤ８Ｔ細胞」というリンパ球の浸潤を抑える
ＰＡＫ遮断剤、例えば「Ｂｉｏ　３０」などのプロポリスや、ビタミンＫ２（や
はり、ＰＡＫ遮断剤）を豊富に含む納豆などで、いわゆる「メタボ」や肥満症を
治療しうる可能性が出てきた。詳しくは、下記を参照されたし。


メタボの原因は脂肪の炎症　薬で治療できるかも？

朝日新聞 (2009年7月27日) より

　メタボリック症候群は、肥満でたまった内臓脂肪に免疫細胞が集まって炎症状
態を起こすことが原因であると、東京大の研究グループがマウスの実験で確かめ
た。糖尿病などの生活習慣病をまねくメタボを免疫を調整する薬で抑えられる可
能性を示した成果で、２６日付の米医学誌ネイチャーメディシン電子版に発表し
た。

　東京大の真鍋一郎特任准教授と西村智特任助教ら循環器内科のグループは、蛍
光色素を注射して内臓の脂肪組織の細胞をそのまま観察できる方法を開発、高脂
肪のエサを与えたマウスの内臓の状態を調べた。

　すると、マウスが太って脂肪細胞が大きくなるにつれ、病原体を攻撃する「Ｃ
Ｄ８Ｔ細胞」というリンパ球の一種が出現し、さらに各種の免疫細胞が集まり、
炎症状態になっていることが確認できた。

　ＣＤ８Ｔ細胞を働かなくしたり、なくしたりしたマウスでは、高脂肪のエサを
与えても、内臓脂肪に免疫細胞が集まる炎症の状態は起こらなかった。さらに、
ＣＤ８Ｔ細胞がないマウスに、この細胞を入れると、脂肪組織に炎症が起きた。

　また、脂肪組織の炎症が起きているマウスに、この細胞への抗体を与えると、
インスリンが効きやすくなり、血糖値が下がった。

　肥満した人の脂肪組織でも同じようなことが起きているのかはまだ不明だが、
グループの永井良三・東京大教授（循環器内科）は「ＣＤ８Ｔ細胞を抑える薬な
どでメタボリック症候群による生活習慣病を治療できる可能性が出てきた」とし
ている。（本多昭彦）

(注)：
実は、パリ大学医学部の研究グループが２００８年に、同様な結果を既に発表して
いる（下記の英文要旨を参照されたし）。

Gastroenterology. 34:1459-69.

Obesity-induced lymphocyte hyperresponsiveness to chemokines: a new mechanism
of Fatty liver inflammation in obese mice.

Bigorgne AE, Bouchet-Delbos L, Naveau S, Dagher I, Prevot S, Durand-Gasselin
I, Couderc J, Valet P, Emilie D, Perlemuter G. (2008)

INSERM, U764, Clamart, France; Univ Paris-Sud, Faculte de Medecine Paris-Sud,
Institut Federatif de Recherche 13, Clamart, France.

BACKGROUND & AIMS: Hepatic lipid retention (steatosis) predisposes hepatitis.
We investigated the mechanisms of lymphocyte homing to fatty liver and
the role of lipopolysaccharide (LPS) in the onset of inflammation in ob/ob
mice.

METHODS: We decreased intestinal bacterial compounds by oral antibiotic
treatment to test the role of endogenous LPS in liver inflammation. Adoptive
transfer of lymphocytes was used to study the respective contributions of
steatosis and lymphocytes to liver inflammation. We tested lymphocyte response
to chemokines by in vitro chemotaxis assays in ob/ob, their lean controls,
and "non-obese ob/ob" mice, generated by controlling caloric intake to distinguish
between the effects of obesity and leptin deficiency.

RESULTS: Antibiotic treatment decreased liver infiltration with CD4(+) T,
CD8(+) T, natural killer (NK)T, B, and NK cells. Adoptive transfer of lymphocytes
from ob/ob or control mice showed that (1) steatosis increased lymphocyte
recruitment to the liver; (2) CD4(+) T, CD8(+) T, and B cells from ob/ob
mice had a greater propensity to migrate specifically to the liver. This
migration was enhanced by LPS. These results were also observed in a model
of high-fat diet-induced obesity. CD4(+) T and B cells were hyperresponsive
to CXCL12 and CXCL13, respectively. Weight normalization in "non-obese ob/ob"
mice decreased liver inflammation, lymphocyte response to chemokines, and
homing to the liver.

CONCLUSIONS: Our study provides the first evidence that liver inflammation
in mice with genetic or diet-induced obesity results from both steatosis
and lymphocyte hyperresponsiveness to chemokines expressed in the liver.
These abnormalities are reversible with weight normalization.

Curcumin could cure both NF2 and TSC (tuberous sclerosis) by "double" punches, knocking out PAK1 and TOR

This year a group at LSU in Louisiana found that Curcumin blocks the Raptor-TOR
(target of rapamycin) interaction which is essential for the activation
of TOR. The kinase TOR is abnormally activated when the tumor suppressors, TSC1 or
TSC2, dysfunction. The rare but formidable tumors associated with TSC (tuberous
sclerosis), where TSC1 or TSC2 dysfunctions, require TOR for their growth.
In animal experiments anti-TOR drugs such as rapamycin could suppress the
growth of TSC tumors, but none of these anti-TOR drugs is available on the
market as yet. However, the natural anti-cancer product "Curcumin" could
be potentially useful for the treatment of TSC, if its bioavailablity is
dramatically improved by either liposome, CD or propolis in the future, as
mentioned before.

Furthermore, two US groups, at Memorial Sloan-Kettering Cancer Center in
New York and MGH in Boston, reported recently that dysfunction of Merlin,
the NF2 gene product, which is the cause of NF2 tumors, also leads to the abnormal
activation of TOR. We found several years ago that Merlin is a direct PAK1
inhibitor (Hirokawa, Y. et al, 2004). It still remains to be clarified whether
the abnormal activation of TOR is through the kinase PAK1, which is abnormally
activated by the dysfunction of Merlin. Nevertheless, this finding suggests
that "Curcumin" could suppress the growth of NF2 tumors by at least two
ways, inactivating both PAK1 and TOR.

In this context, it would be of great interest to note that the tumor suppressor
"FOXO"/Daf-16 in a tiny nematode called C. elegans blocks the expression
of "Raptor", thereby inactivating "TOR". This potentially interesting finding
was reported in 2004 by Don Riddle's group at University of Missouri. Furthermore,
we recently found that the kinase PAK1 inactivates "FOXO" in this worm.

Thus, natural anti-PAK1 products such as Bio 30 and "Curcumin" would inactivate
"TOR" by reactivating "FOXO" at least in this worm. It would be certainly
worth testing whether this remains true with mammals, too.

To be continued

Beevers CS, Chen L, Liu L, Luo Y, Webster NJ, Huang S. (2009).
Curcumin disrupts the Mammalian target of rapamycin-raptor complex.
Cancer Res. 69, 1000-8.

Department of Biochemistry and Molecular Biology, Feist-Weiller Cancer Center,
Louisiana State University Health Sciences Center, Shreveport, Louisiana
71130-3932, USA.

Curcumin (diferuloylmethane), a polyphenol natural product of the plant
Curcuma longa, is undergoing early clinical trials as a novel anticancer
agent. However, the anticancer mechanism of curcumin remains to be elucidated.
Recently, we have shown that curcumin inhibits phosphorylation of p70 S6
kinase 1 (S6K1) and eukaryotic initiation factor 4E (eIF4E) binding protein
1 (4E-BP1), two downstream effector molecules of the mammalian target of
rapamycin complex 1 (mTORC1) in numerous cancer cell lines. This study was
designed to elucidate the underlying mechanism. We observed that curcumin
inhibited mTORC1 signaling not by inhibition of the upstream kinases, such
as insulin-like growth factor 1 receptor (IGF-IR) and phosphoinositide-dependent
kinase 1 (PDK1). Further, we found that curcumin inhibited mTORC1 signaling
independently of protein phosphatase 2A (PP2A) or AMP-activated protein kinase
AMPK-tuberous sclerosis complex (TSC). This is evidenced by the findings
that curcumin was able to inhibit phosphorylation of S6K1 and 4E-BP1 in
the cells pretreated with PP2A inhibitor (okadaic acid) or AMPK inhibitor
(compound C), or in the cells expressing dominant-negative (dn) PP2A, shRNA
to PP2A-A subunit, or dn-AMPKalpha. Curcumin did not alter the TSC1/2 interaction.
Knockout of TSC2 did not affect curcumin inhibition of mTOR signaling.
Finally, we identified that

curcumin was able to dissociate raptor from mTOR, leading to inhibition
of mTORC1 activity. Therefore, our data indicate that curcumin may represent
a new class of mTOR inhibitor.


Lopez-Lago MA, Okada T, Murillo MM, Socci N, Giancotti FG. (2009)
Loss of the tumor suppressor gene NF2, encoding merlin, constitutively activates
integrin-dependent mTORC1 signaling.
Mol Cell Biol. 29, 4235-49.

Cell Biology Program, Sloan-Kettering Institute for Cancer Research, Memorial
Sloan-Kettering Cancer Center, 1275 York Ave., New York, NY 10065, USA.

Integrin signaling promotes, through PAK1, phosphorylation and inactivation
of the tumor suppressor merlin, thus removing a block to mitogenesis in
normal cells. However, the biochemical function of merlin and the effector
pathways critical for the pathogenesis of malignant mesothelioma and other
NF2-related malignancies are not known. We report that integrin-specific
signaling promotes activation of mTORC1 and cap-dependent mRNA translation.

Depletion of merlin rescues mTORC1 signaling in cells deprived of anchorage
to a permissive extracellular matrix, suggesting that integrin signaling
controls mTORC1 through inactivation of merlin.

This signaling pathway controls translation of the cyclin D1 mRNA and, thereby,
cell cycle progression. In addition, it promotes cell survival. Analysis
of a panel of malignant mesothelioma cell lines reveals a strong correlation
between loss of merlin and activation of mTORC1.

Merlin-negative lines are sensitive to the growth-inhibitory effect of rapamycin,
and the expression of recombinant merlin renders them partially resistant
to rapamycin. Conversely, depletion of merlin restores rapamycin sensitivity
in merlin-positive lines. These results indicate that integrin-mediated
adhesion promotes mTORC1 signaling through the inactivation of merlin. Furthermore,
they reveal that merlin-negative mesotheliomas display unregulated mTORC1
signaling and are sensitive to rapamycin, thus providing a preclinical rationale
for prospective, biomarker-driven clinical studies of mTORC1 inhibitors
in these tumors.

"Combo" Therapy of PAK1-dependent Cancers and NF by Two Inexpensive Natural Products, Bio 30 and Curcumin

NF (neurofibromatosis) tumors and 70% of human cancers require the oncogenic
kinase PAK1 for their growth. We have recently shown that Bio 30, the CAPE
(caffeic acid phenethyl ester) -rich extract of NZ (New Zealand) propolis
almost completely suppresses the growth of NF tumors, glioma, pancreatic
and breast cancer xenografts in mice. Another natural product called curcumin
was also known to block this kinase somehow. This summer Feng Li's group
at China Medical School reported that curcumin inactivates PAK1, at least
in part by down-regulating the Tyr-kinase ErbB2/HER2 (for detail, see the
abstract below). Several years ago we have shown that both ErbB1 (EGF receptor)
and ErbB2 are required for PAK1 activation (Hong He et al, 2001).

The major problem of curcumin alone for clinical application is its poor
bioavailability (mainly due to its water-insolubility) as previously discussed.
In the future, liposome or CD (cyclodextrin) could be used to solubilize
curcumin for clinical application. Interestingly, CAPE alone shares the same
problem. However, if CAPE is taken with several other anti-cancer polyphenols
in CAPE-based propolis extracts such as Bio 30, both its bioavailability
and anti-cancer potential are dramatically (over 600 times) improved (Maria
Demestre et al, 2009). Considering this finding, it would be worth to combine
curcumin and Bio 30 not only for improving the bioavailability of curcumin,
but also for creating its potential synergy with CAPE and several other
anti-cancer ingredients such as pinocembrin (PIN) in Bio 30.


Cancer Biol Ther. 2009 Jul 13;8(14).

Curcumin suppresses proliferation and invasion in human gastric cancer cells
by downregulation of PAK1 activity and cyclin D1 expression.

Cai XZ, Wang J, Li XD, Wang GL, Liu FN, Cheng MS, Li F.

Department of Cell Biology, Key Laboratory of Cell Biology of Ministry of
Public Health of China, China Medical University, Shenyang, China.

Curcumin (diferuloylmethane), is a natural chemopreventive agent known to
inhibit the proliferation of several cancer cell lines. It has been previously
demonstrated that curcumin is a potent inhibitor of EGF-receptor (EGFR)
tyrosine kinase, but its inhibitive effect on PAK1, a downstream protein
of EGFR, has not been defined. In this paper

we found that curcumin repressed the expression of HER2 and inhibited the
kinase activity of PAK1 without affecting its expression. Silencing HER2
in gastric cancer cells showed that even if PAK1 activity was transiently
strengthened by EGF, curcumin still had a strong inhibitive effect. It should
be emphasized that kinase assay in vitro showed that curcumin could act as
an ATP-competitive inhibitor, which was supported by computer-aided molecular
modeling. Curcumin also downregulated the mRNA and the protein expression
of cyclin D1 and suppressed transition of the cells from G(1) to S phase.
Therefore, curcumin inhibited the proliferation and invasion of gastric
cancer cells. Overall, these results provided novel insights into the mechanisms
of curcumin inhibition of gastric cancer cell growth and potential therapeutic
strategies for gastric cancer.

OSU-03012, the "Synthetic" Anti-PAK1 Drug, Blocks the Growth of NF2 Tumor Xenografts in Mice.

In 2007 a group led by Matthew Ringel at OSU (Ohio State University) found
that a synthetic compound called OSU-03012 selectively inhibits PAK1 in
vitro with IC50 around 1 micro M, and blocks the growth of several thyroid
cancer cell lines. In other words thyroid cancer is among PAK1-depenent
cancers. This compound is a derivative of the cyclooxygenese-2 inhibitor
"celecoxib", and was initially developed as a PDK1 inhibitor by Ching-Shih Chen's
group at OSU in 2004. It eventually inactivates another kinase AKT
by inhibiting the kinase PDK1. However, since it inhibits the growth of
PAK1-dependent (and AKT-independent) cancer cell lines such as PC-3 (PTEN-deficient
prostate cancer) and pancreatic cancer cells, it had been suggested that
OSU-03012 might block the kinase PAK1. In 2009 OSU-03012 (200 mg/kg, daily)
was found by Long-Sheng Chang's group at OSU to suppress the growth of NF2-deficient tumors
(Schwannoma) xenografts in mice by more than 50% (for detail, see the abstract below).

It should be worth to point it out that like ARC (Artepillin C) from Brazilian
green propolis, OSU-03012 is among the first single chemical drugs that
were shown to block the growth of NF2 tumors in vivo, although Bio 30
(100 mg/kg, twice a week) a mixture of several anti-cancer ingredients
including CAPE, was proven to block completely the growth of NF2 tumors in vivo, and is inexpensively available
on the market. If OSU-03012 is successful in the time and money-consuming
clinical trials for these PAK1-dependent cancers and NF tumors, this "synthetic"
compound would become available on the market in a decade or so... Hope
(unlike FK228) this drug can pass BBB (blood brain barrier) so that it works
on formidable brain tumors such as glioma and NF2 tumors.

Eur J Cancer. 2009 Jun; 45(9):1709-20

Growth inhibitory and anti-tumour activities of OSU-03012, a novel PDK-1
inhibitor, on vestibular schwannoma and malignant schwannoma cells.

Lee TX, Packer MD, Huang J, Akhmametyeva EM, Kulp SK, Chen CS, Giovannini
M, Jacob A, Welling DB, Chang LS.

Department of Otolaryngology, The Ohio State University College of Medicine,
Center for Childhood Cancer, The Research Institute at Nationwide Children's
Hospital, Columbus, OH, USA.

BACKGROUND: Vestibular schwannomas (VS) frequently express high levels of
activated AKT. Small-molecule inhibitors of AKT signalling may have therapeutic
potential in suppressing the growth of benign VS and malignant schwannomas.
METHOD: Primary VS and Schwann cells, human malignant schwannoma HMS-97
cells and mouse Nf2(-/-) Schwann cells and schwannoma cells were prepared
to investigate the growth inhibitory and anti-tumour activities of OSU-03012,
a celecoxib-derived small-molecule inhibitor of phosphoinositide-dependent
kinase-1. Cell proliferation assays, apoptosis, Western blot, in vivo xenograft
analysis using SCID mice and immunohistochemistry were performed.

RESULTS: OSU-03012 inhibited cell proliferation more effectively in both
VS and HMS-97 cells than in normal human Schwann cells. The IC5) of OSU-03012
at 48h was approximately 3 micro M for VS cells and HMS-97 cells, compared
with the IC(50) of greater than 12 micro M for human (normal) Schwann cells.
Similarly, mouse Nf2(-/-) schwannoma and Nf2(-/-) Schwann cells were more
sensitive to growth inhibition by OSU-03012 than wild-type mouse Schwann
cells and mouse schwannoma cells established from transgenic mice carrying
the NF2 promoter-driven SV40 T-antigen gene. Like VS cells, malignant schwannoma
HMS-97 cells expressed high levels of activated AKT. OSU-03012 induced apoptosis
in both VS and HMS-97 cells and caused a marked reduction of AKT phosphorylation
at both the Ser-308 and Thr-473 sites in a dose-dependent manner.

In vivo xenograft analysis showed that OSU-03012 was well tolerated and
inhibited the growth of HMS-97 schwannoma xenografts by 55% after 9 weeks
of oral treatment. The anti-tumour activity correlated with reduced AKT
phosphorylation.

CONCLUSION: OSU-03012 is a potential chemotherapeutic agent for VS and malignant
schwannomas.

PAK1-LC8 (Dynein Light Chain) Interaction Required for Nuclear Import of PAK1.

PAK1 is an oncogenic Ser/Thr kinase which is activated by several cytoplasmic
proteins such as GTPases (Rac and CDC42), SH3 adaptor proteins (PIX and
NCK), and Tyr-kinase (ETK). This kinase is required for the growth of more
than 70% of human cancers, including breast and prostate cancers, as well
as formidable brain tumors such as glioma and NF (neurofibromatosis) tumors.

Recently John Williams' group at Thomas Jefferson University in Philadelphia
found that Dynein Light Chain (LC8) also binds residues 212-222 of PAK1
which is adjacent to the essential NLS (nuclear localization site, residues
243-245), and their interaction leads to the dimerization of PAK1 through
NLS and is required for the EGF-induced nuclear import of the cytoplasmic
PAK1 in MCF-7, an estrogen-dependent breast cancer cell line. Since PAK1
phosphorylates the nuclear estrogen receptor (ER) at Ser 305 for the ER
activation which leads to the estrogen-dependent malignant growth of this
cell line, LC8 appears to contribute to the oncogenicity of PAK1 by nuclear
import.


PLoS One. 2009 Jun 26;4(6):e6025

Interaction with LC8 is required for Pak1 nuclear import and is indispensable
for zebrafish development.

Lightcap CM, Kari G, Arias-Romero LE, Chernoff J, Rodeck U, Williams, John
C.

Department of Biochemistry and Molecular Biology, Thomas Jefferson University,
Philadelphia, PA, USA.

Pak1 is a serine/threonine kinase implicated in regulation of cell motility
and survival and in malignant transformation of mammary epithelial cells.
In addition, the dynein light chain, LC8, has been described to cooperate
with Pak1 in malignant transformation of breast cancer cells. Pak1 itself
may aid breast cancer development by phosphorylating nuclear proteins, including
estrogen receptor alpha. Recently, we showed that the LC8 binding site on
Pak1 is adjacent to the nuclear localization sequence (NLS) required for
Pak1 nuclear import.

Here, we demonstrate that the LC8-Pak1 interaction is necessary for epidermal
growth factor (EGF)-induced nuclear import of Pak1 in MCF-7 cells, and that
this event is contingent upon LC8-mediated Pak1 dimerization. In contrast,
Pak2, which lacks an LC8 binding site but contains a nuclear localization
sequence identical to that in Pak1, remains cytoplasmic upon EGF stimulation
of MCF-7 cells.

Furthermore, we show that severe developmental defects in zebrafish embryos
caused by morpholino injections targeting Pak are partially rescued by co-injection
of wild-type human Pak1, but not by co-injection of mutant Pak1 mRNA disrupting
either the LC8 binding or the NLS site.

Collectively, these results suggest that LC8 facilitates nuclear import
of Pak1 and that this function is indispensable during vertebrate development.