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Translocating Lactobacillus torments tumors

via tryptophan catabolism
Márcia S. Pereira1 and Martin A. Kriegel1,2,3,4,*
1Department of Translational Rheumatology and Immunology, Institute of Musculoskeletal Medicine, University of Münster, 48149 Münster,
Germany
2Section of Rheumatology and Clinical Immunology, Department of Internal Medicine, University Hospital Münster, 48149 Münster, Germany
3Cells in Motion Interfaculty Centre, University of Münster, 48149 Münster, Germany
4Department of Immunobiology, Yale University School of Medicine, New Haven, CT 06511, USA

*Correspondence: martin.kriegel@ukmuenster.de
https://doi.org/10.1016/j.cell.2023.03.022

Variability in the efficacy of immune checkpoint inhibitors in cancer patients is associated with the human gut
microbiota. However, detailed mechanisms are unclear. In this issue of Cell, Bender et al. uncovered that a
probiotic Lactobacillus strain translocates into murine tumors to enhance immunotherapy via the tryptophan
metabolite indole-3-aldehyde (I3A).

The gut microbiota profoundly influences
immune cell development, differentiation,
and function. These processes coevolved
with the microbiota over millennia but
have been disrupted over the last decades
due to modern lifestyle. Dysregulated
host-microbiota interactions are increas-
ingly linked to a variety of immunologic dis-
eases,1 malignancies, and altered anti-
tumor responses.2 Antitumor immunity is
unleashed by inhibition of so-called im-
mune checkpoints, which improves can-
cer patient survival and has revolutionized
cancer immunotherapy. One of these im-
mune checkpoint inhibitors (ICI) blocks
programmed death-ligand 1 (PD-L1) -
PD-1 interactions, thereby allowing other-
wise restrained cytotoxic T cells (CTLs) to
attack tumor cells. These responses, how-
ever, vary among and within patient co-
horts and types of cancers. One reason
for this variability in ICI responsiveness is
the human gut microorganisms and their
genetic material referred to as the gut mi-
crobiome.2 Accordingly, probiotic gut bac-
teria have been shown to modulate ICI re-
sponses in patients and corresponding
animal models. However, the mechanisms
of how resident or exogenously adminis-
tered bacteria contribute to ICI therapies
were poorly understood. Publishing in
this issue of Cell, Bender et al.3 dissected
one of these mechanisms and could link
a microbially derived metabolite to ICI
responsiveness in cancer patient cohorts.
Bender et al.3 tested if and how a probi-
otic strain of Limosilactobacillus reuteri
(L. reuteri) enhances anti-PD-L1 or anti-
cytotoxic T-lymphocyte associated pro-
tein 4 (CTLA-4) ICI therapy in a melanoma
mouse model induced by injecting B16
melanoma cells subcutaneously in the
hind flank. Oral gavage of L. reuteri sup-
pressed the growth of melanoma with
and without ICI in a CD8+ T cell- and inter-
feron (IFN)-g-dependent manner. Orally
administered L. reuteri also provided
potent anticancer effects in three addi-
tional solid tumor models. Importantly,
small intestinal L. reuteri was found to
translocate to, and to persist within, the tu-
mors. Translocation promoted the intratu-
moral expansion of IFN-g+ and perforin+
CTLs with antitumor activity. Adoptive
transfer studies confirmed that CD8+
T cells are sufficient for antitumor immu-
nity. Cytotoxic T cell activity required the
intratumoral release of the tryptophan
metabolite indole-3-aldehyde (I3A) from
L. reuteri. I3A subsequently activated the
aryl hydrocarbon receptor (AhR) in CD8+
T cells but was not required in other im-
mune cells as revealed in vivo by selective
genetic deletion of AhR in cellular subsets.
A tryptophan-enriched diet or I3A supple-
mentation also increased AhR activity in
the tumor and suppressed tumor growth
in mice. Similarly, intratumoral injection
of I3A, but not kynurenine, suppressed

Cell 186, April 27, 2023 ª 2023 Elsevier Inc. 1821

 ll
Figure 1. Probiotic Lactobacillus enhances cancer immunotherapy by translocation intratumorally
Limosilactobacillus reuteri activates an indole-3-aldehyde (I3A)-aryl hydrocarbon receptor (AhR)-CD8+ T cell axis intratumorally after translocation from the small
intestine into the tumor microenvironment. The AhR agonist I3A can also be derived from a tryptophan-rich diet and activates cytotoxic CD8+ T cells that secrete
IFN-g. This antitumor effect enhances immune checkpoint inhibitor therapy with anti-PD-L1 antibodies.
melanoma growth, mimicking the effects
of translocated L. reuteri. Finally, Bender
et al.3 extended the preclinical findings to
cancer patient cohorts that showed vari-
able outcomes to ICI therapy. They found
that patients responding to ICI carried
higher levels of I3A in the serum, whereas
levels of the endogenous AhR ligand
kynurenine were not different between
responders and non-responders. Interest-
ingly, another microbially derived trypto-
phan metabolite produced distantly
within the gut was recently shown to
enhance chemotherapy in pancreatic
cancer.4 Furthermore, another recent
study demonstrated also that translo-
cated microbiota support ICI antitumor
immunity.5
By taking all findings together, Bender
et al.3 delineate a mechanism whereby a
probiotic bacterial strain translocates
from the small intestine into the tumor
microenvironment of solid cancers in ani-
mal models and enhances ICI therapy via
an I3A-AhR-CD8+ CTL axis (Figure 1).
This study greatly enhances our under-
standing of probiotic-mediated antitumor
immune responses but also raises new
questions. How does L. reuteri cross the
gut barrier in the first place? Bender
et al.3 tested various alterations in gut
epithelial, vascular, and lymphatic bar-
riers but did not find major phenotypic
or functional abnormalities. However,
L. reuteri could more subtly promote bar-
1822 Cell 186, April 27, 2023
rier dysfunctions to escape into extrain-
testinal tissues. Alternatively, it might
passively enter the host given the rapid
translocation characterized by Bender
et al.3 Further, Bender et al.3 showed
that translocation to tumors, but not sec-
ondary lymphoid organs, is required for
L. reuteri-mediated cancer suppression.
Which route does L. reuteri take to reach
the tumor? Does it travel attached or
within chemotactically attracted immune
cells or independently of host cells?
Where and how does it persist in the tu-
mor microenvironment without being
eliminated by the host immune system?
Does L. reuteri interact with tumor-type
specific intracellular bacteria that were
recently characterized?6 L. reuteri could
be taken up by circulating immune cells
that are attracted to the tumor microenvi-
ronment by chemotactic signals. In that
case, however, immune evasive strate-
gies need to be exploited by L. reuteri to
avoid destruction of the ‘‘trojan horse’’
by a host immune response. Recent
studies on long-term colonization of
L. reuteri and unrelated Enterococcus gal-
linarum in autoimmune models support
within-host evolution influencing translo-
cation and immune evasion.7 The rapid
translocation and antitumor effects of
probiotic L. reuteri are likely not affected
by within-host evolution but this phenom-
enon may impact endogenously colo-
nizing L. reuteri strains in cancer patients.
Previews
Antitumor immunity after ICI therapy
is frequently associated with immune-
related adverse events,8 suggesting
that the two processes are linked.
L. reuteri is known to translocate to
the liver and secondary lymphoid organs
in autoimmune susceptible models.9,10
Translocation in this setting exacerbates
systemic9 and organ-specific10 autoim-
mune pathology. Bender et al.3 did not
see signs of systemic autoimmunity
serologically, but it remains to be deter-
mined if probiotic therapy with L. reuteri
may increase the risk for ICI-associated
autoimmune pathology. In addition,
sepsis with probiotic strains have been
documented in highly immunocompro-
mised states,11 which could also occur
in cancer patients receiving concomitant
chemotherapy.
Still, the possibility to treat cancer pa-
tients with probiotics or their metabolites
is an exciting new avenue in cancer immu-
notherapy. Personalized approaches may
be needed given that probiotics do not
colonize every human subject equally,12
but Bender et al.3 also provide evidence
for circumventing live biotherapeutic
therapies by supplementing mediators of
antitumor immunity (in this case I3A) or
by dietary approaches that enrich them.
Multiple approaches thus exist to harness
the power of microbiota-based therapies
in oncology. The current study provides
insights into mechanisms that may lead

Previews
to additional targeted therapies within the
microbiome or the host. In either case,
how viable bacteria translocate to the tu-
mor microenvironment warrants further
study to better understand how immune-
related adverse events and autoimmunity
might arise. More broadly, context- and
strain-specific effects of the gut micro-
biota on extraintestinal immune re-
sponses as illuminated by Bender et al.3
underscore our interconnectedness with
the microbial world.
DECLARATION OF INTERESTS
M.A.K. received salary, consulting fees, honoraria,
or research funds from Eligo Biosciences, Enter-
ome, Roche, Novartis, Bristol–Meyers Squibb,
AbbVie, GlaxoSmithKline, MSD Sharpe & Dohme,
and Cell Applications, and holds a patent on the
use of microbiota manipulations to treat immune-
mediated diseases. M.S.P. declares no competing
interests.
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