A specific bacteria-infecting virus, bacteriophage, found in gut microflora, augments anti-tumor T cell immunity

Molecular mimicry between microbial and host's antigens could contribute to autoimmunity but also to the protection against tumors through epitope cross-reactivity. A new study in journal Science indicates that those cross-reactive epitopes could come from viruses that infect endogenous microbial species. 

In this study the authors made a surprising observation that only certain Enterococcus hirae microbial strains (E. hirae 13144 or IGR11) augmented anti-cancer effect in experimental cancer model.

 

 

Next, the authors showed that this biological activity was linked to one dominant epitope, TSLARFANI, derived from TMP protein that originated in 39.2-kb prophage only in those specific E. hirae strains. Mice immunized with heat-inactivated E. hirae 13144 strain, or peptide TSLARFANI, or irrelevant E.coli engineered to express TMP, all augmented anti-cancer effect. 

 

 

Mechanistically, the authors showed that epitope, GSLARFRNI, derived from cancer cells used in these experiments, was recognized by the same CD8 T cells which labeled with TSLARFANI epitope tetramers confirming cross-reactivity between these 2 epitopes.

 

 

In summary, this study suggests that microbiota and bacteriophages they carry represent new modality in fight against cancer. In this study overall anti-tumor effect is modest but we need to take into account that this is an effect of just one cross-reactive epitope in one type of MHC inbred mice, and it is likely that many other epitopes will be involved in outbred species such as humans. However, it is still extremely hard to do such analysis in humans in real world scenario due to lack of exact knowledge about human microbiota strains and poor reliability and performances of available bioinformatics approaches. However, once the mechanistic principles underlying anti-cancer effects are uncovered and accepted, then it is much easier to move the field forward.

posted by David Usharauli

Tumor elimination requires simultaneous expression of both class I and II neo-epitopes

The most tumors express mutant epitopes that could be detected by T cells. According to current paradigm, CD4+ T cells provides help to CD8+ T cells that in turn attack tumors. As tumor cells ordinarily express class I recognized by CD8+ T cells but not class II molecules recognized by CD4+ T cells, primary focus on CD8+ T cell epitopes made a lot of sense. But what about CD4+ T cell 'help' to CD8 T cells? 

Indeed, a new 'classically-done' immunology study from Robert Schreiber's lab clearly showed that irrespective class II expression, tumor cells must express both CD8+ and CD4+ T cell neo-epitopes to achieve efficient local tumor control following immunotherapy.

As a starting point, they used nonimmunogenic oncogene-driven KP9025 sarcoma cells (KP), which lack mutational neoantigens. Next they re-expressed in KP cells 2 mutant epitopes, one for class I, mLAMA4, and another for class II, mITGB1 (identified using a hidden Markov model (HMM)-based MHC binding predictor the authors claim is better than other available algorithms). 

 

A mutant but not wild-type version of ITGB1 was detected by CD4+ TILs.

  

Next, the authors showed that only KP tumors expressing both neo-epitopes but not single expressors, could be eliminated by T cells following immunotherapy.

  

As expected, presence of CD4+ T cell epitope enhanced CD8+ T cell response.

Interestingly, both class I and II  neo-epitopes must be expressed by the same tumor to mediate protection when used as immunized agents (mixing of single expressor tumors was not enough).

And notably, expression of both class I and II neo-epitopes were necessary to mediate efficient local tumor control (single expressor tumors were resistant against CD8+ or CD4+  T cells)

In summary, this is a simple, easy to follow experments that indicate the authors' thought process.  It shows that CD4+ T cells 'help' to CD8+ T cells are required both at priming and as well as at effector stage. It is not clear if it is simply a quantitative or rather qualitative issue. It is not known either whether CD4+ T cells do something directly against tumor beyond simply helping CD8+ T cells here. 

posted by David Usharauli

β-synuclein specific CD4+ T cells invade brain grey matter to cause array of MS-like symptoms

β-synuclein specific CD4+ T cells invade #brain grey matter and cause array of MS-like #symptoms in a rat model of human #MultipleSclerosis. Interestingly, the most useful distilled data points were in suppl materials. #immunology #autoimmunedisease https://t.co/NbIt5ijxuR

— David Usharauli (@3DiMMUNE) March 5, 2019

highlights from paper
2. Such exclusive invasion of brain grey matter were primarily correlated by pattern of #antigen (β-synuclein) expression. #autoimmunedisease #immunology pic.twitter.com/1r9ROgtTDV

— David Usharauli (@3DiMMUNE) March 5, 2019

Some thoughts: here the authors have used either WT or transgenic rat expressing β-synuclein-specific T cells. Diseases were induced either following direct immunization of transgenic rats or adoptive transfer of activated β-#synuclein-specific T cell blasts. #immunology

— David Usharauli (@3DiMMUNE) March 5, 2019

Con't: immunologists have fairly good idea about processes that represent immune response but at presently it is still unknown how immune response is #initiated naturally or how it is #stopped. This is Achilles heels of current experimental models in #immunology.

— David Usharauli (@3DiMMUNE) March 5, 2019

An antigen from gut commensal Bacteroides thetaiotaomicron (B. theta) is recognized by Foxp3+ Tregs

1. B.theta-specific T cells when transferred into #antibiotic-treated and B.theta #colonized T cell-deficient mice (#RAG1-KO) develop into #Foxp3+ #Tregs as well as effector T cells. It is not clear if donor tg T cells were on RAG1-KO background as well. Host mice stay healthy. pic.twitter.com/TD0pnnxgAR

— David Usharauli (@3DiMMUNE) February 9, 2019

3. The authors were able to identify a #specific #epitope in B.theta recognized by B.theta-specific T cells. The #sequence, EEFNLPTTNGGHAT, were mapped in BT4295 predicted to be a SusE/SusF #lipoprotein in outer #membrane (OM). pic.twitter.com/rH5cQHQcVN

— David Usharauli (@3DiMMUNE) February 9, 2019

In #summary, this study showed that certain gut #commensal #microbiota species, in this case, #Bacteroides thetaiotaomicron (B. theta) are recognized by #Tregs and even induce #effector T cell differentiation though the host stays healthy because Tregs regulate effector T cells.

— David Usharauli (@3DiMMUNE) February 9, 2019

Another point is whether #microbiota-specific Tregs are #denovo induced or #amplified from existing ones. I personally think it is a latter. Most #Tregs develop from #CD25-negative #precursors poised for Foxp3 expression & such precursors are all #thymus derived, in my opinion.

— David Usharauli (@3DiMMUNE) February 9, 2019

Hypocretin-specific T cells mediate autoimmune sleeping disorder narcolepsy

Narcolepsy is a sleeping disorder characterized by "excessive daytime sleepiness, cataplexy, hypnagogic hallucinations and sleep paralysis". In recent years several studies provided evidence suggested that etiology of narcolepsy could be autoimmune in nature. Such conclusion has become more mainstream especially following observed relationship between 2009 pandemic flu vaccine, Pandemrix (from GSK), vaccination and development of sleeping disorder in a subset of vaccine recipients. In one study, it was found that Pandemrix but not Focetria flu vaccine (they differ in only one amino acid from each other) caused anti-flu antibody production that cross-reacted with hypocretin receptor.

Now, new study in Nature went further and showed that individuals suffering from narcolepsy, but not healthy controls, harbor hypocretin-specific T cells.

Antigen-specific T cells are rare so to reveal their presence the authors first non-specifically amplify T cells from PBMCs and after considerable expansion tested on hypocretin peptide pool pulsed autologous B cells. Majority of individuals with narcolepsy showed reactivity in this assay.

Single cell analysis showed that hypocretin-specific T cells consist of several clones (up to 30 different clones).       

Interestingly, most of T cells specific for hypocretin were HLA-DR restricted, rather than HLA-DQ as earlier association studies would have predicted.

Also, notably, hypocretin-specific T cells did not reacted with autologous B cells (or monocytes) pulsed with hypocretin protein suggested that this in vitro pulsing assay could not recapitulate in vivo hypocretin protein processing pathway. 

Of note, the authors showed that hypocretin-specific T cells did not cross-react with pandemic flu peptide pool or vaccine (influvac). However, they did not use Pandemrix vaccine here so these results are not conclusive. 

In summary, this study showed that narcolepsy could indeed be a bona fide autoimmune disease. As in many other autoimmune diseases exact molecular events that initiate them are yet to be discovered.

posted by David Usharauli 

Cross-reactivity between microbial-derived antigens and tumor neoantigens correlates with long-term survival

This is a very interesting paper published in Nature few months back. In this study the authors wanted to uncover immune correlates of long-term (>10 yrs) survival from pancreatic ductal adenocarcinoma that normally account for less than 2% of all patients. 

First they found that "patients with both the highest predicted neoantigen number and either the greatest CD3+CD8+, or polyclonal T-cell repertoire, but neither alone, exhibited the longest survival". It suggested that tumor neoantigen-derived epitope quality rather than simply quantity correlated with survival. 

More importantly, bioinformatics analysis of similarities (mimicry or cross-reactivity or poly-reactivity) and neoantigen fitness modelling between tumor neoatigens and microbial-derived antigens significantly stratified short- and long-term survivors independent of confounding factors and adjuvant chemotherapy. 

If these data will  hold true in other cancer settings, it will append how immunotherapy is applied to treat cancer patients. While this finding is potentially hugely important for immunotherapy, the authors tried not to make too big a statement about it (I would imagine it is a result of a typical reviewers conservatism), writing that "This hypothesis does not assume any associations between pre-existing antimicrobial immunity and survival, but rather aims to develop a strategy to identify candidate neoantigens based on defined immunogenic pathogen-derived epitopes" and in other place "Our results do not indicate causal associations of pre-existing microbial and anti-tumour immunity in LTSs [long-term survivors]. Instead, our data suggest that embedding microbial homology in the context of our neoantigen quality model can help to create an effective surrogate for immunogenic neoantigens." 

posted by David Usharauli

Receptor-ligand specific labeling of immune cell interactions

This week journal Nature published new study in immunology that could be best described as a method paper. I personally don't understand the value of this paper been in Nature. Only positive characteristic I see in it is that experiments reported are done in a classical, cellular immunology "fashion" and very easy to follow and understand. Lets examine.

The new method, called LIPSTIC, which this study reported is about labeling receptor-ligand pair with naturally occurring enzyme, the Staphylococcus aureus transpeptidase sortase A (SrtA). SrtA appears to covalently transfers a substrate containing motif ‘LPXTG’ to a nearby oligoglycine (G5). Basically, receptor is genetically fused with SrtA and ligand is fused with G5 and when they interact, SrtA catalyses the transfer of the substrate onto the G5-tagged receptor. This transfer can be visualized by attaching to the substrate small labels such as biotin or a fluorophore.  

First the authors showed in vitro and ex vivo that the substrate transfer was specific to fused receptor-ligand pair and did not occur when enzyme was inactive or fused to unrelated receptor. Most of the reported experiments were done using CD40L/CD40 pair (CD40 signaling is relevant for DCs and CD8 T cell activation by CD40L expressing CD4 T cells).

Receptor-ligand specificity was maintained in vivo as well. Because CD40L upregulation on CD4 T cells depends on antigen-specific interactions, substrate transfer were restricted to those CD40+ DCs that were pulsed with cognate peptide (OVA).

However, this antigen-specific CD40L/CD40 interaction was maintained only for initial 10h-24h period. When OVA-specific T cells were left with DCs for longer period (48h) then even DCs pulsed with irrelevant peptide (LCMV peptide) got labeled with substrate. 

It is not clear what are the biological consequences of such non-specific T/DCs interactions. Are these two different DCs activated antigen or non-antigen specific manner somehow different with the regard of activation of CD8 T cells? We don't know. The problem with such model is that if activated CD4 T cells expressing CD40L can interact with CD40+ DCs and activate it (license it, to use polly matzinger's words) then we should expect that body should harbor only activated DCs because body constantly contains some number of CD40L+ activated CD4 T cells specific for all kind of antigens. It is possible high number of T cells transferred in these experiments created an artificial outcome.

In summary, this study showed new method how to label receptor-ligand pair in vivo. However, overall relevance of this method is not clear at present.

posted by David Usharauli

Engineering T cells to cellular factories with synthetic Notch receptors

This week journal Cell published new article from Wendell Lim's lab at UCSF (also a founder of Cell Design Labs) that reads like a science fiction story. It was a continuation of previous work that focused on developing customized molecular architecture based on Notch core regulatory domain attached to synthetic extracellular recognition and intracellular transcriptional domains (SynNotch). By changing extracellular and intracellular domains one can design T cells producing molecule of interest upon engagement with specific ligand. 

For this new paper, the authors managed to transform T cells into cellular factories which upon specific SynNotch stimulation were making and expressing (a) cytokines (b) checkpoint inhibitors (c) bi-specific antibodies (d) CARs (e) transcriptional factors (f) lytic granules.

Basically, T cells are transfected with vector containing SynNotch module linked to promoter encoding molecule of interest. The most of the experiments were done in vitro. One in vivo experiment the authors put in paper was similar in overall design to one previously reported. In short, NSG mice were injected with K562 tumor expressing GFP + CD19 or only CD19. Afterwards T cells containing SynNotch module designed to recognize GFP were infused. Upon GFP recognition these T cells could start expressing soluble Blinatumomab, a-CD19/CD3 BiTE molecule that in turn can engage conventional TCR and produce T cell activation. Indeed, the authors showed that only tumor cells expressing both GFP and CD19 were efficiently controlled by engineered T cells (though its is not clear why soluble BiTE antibodies would not diffuse freely and engage single CD19+ tumors as well).    

While all these results look very impressive, it nonetheless lack some of the critical elements that are required to properly analyze beneficial effect of SynNotch. For example, in vivo experiments were done in mice that does not express tumor antigens naturally (anti-CD19 here was human in origin). So we can't tell how such T cells would behave in "human-like" environment. Similarly, in vitro experiments with SynNotch-anti-HER2 expressing T cells are of dubious value without testing it at least in humanized mouse models.

David Usharauli

Genomic alterations in IFN-γ pathway underlie resistance to anti-CTLA-4 (Yervoy) therapy

This week journal Cell published a short "reverse translational" study conducted on small cohort of patients non-responsive to immunotherapy with anti-CTLA-4 antibody (ipilimumab, Yervoy) that  showed that genomic alterations in IFN-γ pathway in non-responder patients could underlie their resistance to immunotherapy.  

For this study the authors compared 12 patients who did not respond to ipilimumab therapy (non-responders) and 4 patients who did respond to ipilimumab therapy (responders). They found that "tumor samples from non-responders were found to have significantly more somatic mutations, including copy-number alterations (CNAs) and single-nucleotide variants (SNVs) of the IFN-γ pathway genes".

Next, the authors showed that primary melanoma cell lines derived from anti-CTLA-4 responder or non-responder patients could be differentiated based on their in vitro sensitivity to IFN-γ.

Next, they showed quite bizarre experiment. By knocking down IFN-γ receptor in mouse B16 melanoma cells the authors showed that these cell line became less sensitive to IFN-γ in vitro. Not sure about logic behind these experiment.  

Finally, the authors showed that B16 melanoma cells deficient for IFN-γ signaling and transplanted into WT mice were less sensitive to anti-CTLA-4 therapy.

In summary, this small cohort study suggests that screening of melanoma patients for genomic alterations in IFN-γ pathway could be useful in better predicting therapeutic outcome for this checkpoint inhibitor therapy.

David Usharauli

Tumor-induced T cell dysfunctions are antigen-specific and expressed early on

This week Immunity published new study showing that CD8 T cell dysfunctions frequently observed in tumors is induced (a) early on (between 1-3 weeks), (b) it is antigen-specific (driven by chronic antigenic stimulation) and (c) it is irreversible even when checkpoint inhibitors are applied.

For this paper the authors have used tamoxifen-inducible, autochthonous [indigenous] liver cancer model (ASTxCre-ER^T2; AST = Albumin-floxStop-SV40 large T antigen, Tag). In this model tamoxifen application induces Tag expression that inactivates two central tumor suppressor proteins, retinoblastoma protein and the p53. Analysis of Tag-specific CD8 T cells at weeks 1 or 4 showed that already by week 1 tumor antigen-specific CD8 T effector cells display signs of "exhaustion" phenotype, that was further exaggerated by week 4 (up-regulation of PD-1, 2B4, LAG3, TIM-3 and failure to express cytokines).

Furthermore, when such tumor "educated" CD8 T cells were transferred into tumor-free hosts and challenged with live Tag+ infection, only week 1 CD8 T cells could respond. Week 4 T cells were irreversibly dysfunctional. Week 4 tumor antigen-specific CD8 T cells could not be revived even with checkpoint anti-PD-1 antibody.   

Finally, the authors showed that tumor induced T cell dysfunction was antigen-specific because presence of tumor did not affect non-specific CD8 T cell functionality (Tag-specific vs. OVA-specific T cells).

In summary, this study indicates that tumors could imprint irreversible "epigenetic" effector dysfunction on tumor antigen-specific T cells early on. The results in this study have important implications for clinical immunotherapy. We need to recognize that even with recent progress in application of checkpoint inhibitors for tumor management success is not guaranteed.

One drawback of this study is that the authors did not determine relationship between "antigen dosage" and CD8 T cell dysfunction. Albumin could drive huge amount of tumor antigen expression and the speed of tumor formation in these mice is clear example for such rapid "non-physiological" model. Next step would be to design tumor models where tumor specific antigen expression [dosage] could be regulated as well.

David Usharauli

Harnessing donor TCR specificity for cancer immunotherapy

Cancerous tissues harbor protein mutations that can be recognized by immune system as neoantigens. However, when tumor progresses it indicates that either (A) patient lacks T cells with adequate affinity to tumor neoantigens or (B) tumor environment actively suppresses immune response [or both]. 

For example, novel drug class of checkpoint inhibitors targeting CTLA4 and PD1/PD-L1 inhibitory circuits operating in T cells (Keytruda, Opdivo, Yervoy, Tecentriq) work on option B by modulating tumor suppressive micro-environment.

Another approach obviously would be an option A by using engineered T cells expressing tumor specific T cell receptors. Ideally, patient's own T cells can be expanded and re-infused back to attack tumor cells. But, more likely, patient will lack T cells with  adequate affinity to tumor neoantigens due to TCR editing. 

To overcome this limitation, new study published in journal Science suggested to use instead tumor-specific TCRs harvested from healthy donors. Here, the authors led by T cell expert Ton Schumacher, showed that HLA-matched healthy donors contain T cells with sufficient affinity and specificity to recipient's tumor neoantigens (of note, Ton Schumacher is also affiliated with biotech company Kite Pharma).

This study focused on HLA-A*02:01-restricted neoantigens from stage IV melanoma patients. Neoantigens were identified with whole-exome and RNA sequencing and selected for further analysis based on high predicted binding affinity to HLA-A*02:01. Autologous monocyte-derived dendritic cells transfected with mRNA encoding the candidate epitopes and cultured with healthy donor T cells. All 4 healthy donor  T cells specifically detected mutated tumor neoantigens with greater sensitivity.

Tumor neoantigen-specific T cell response was confirmed in epitope pulse experiment using WT or mut epitopes.

Furthermore, when donor T cell derived TCRs were re-introduced by gene transfer, resulting T cells were specific to patient's mut neoantigen and did not recognize, for example, 3rd party tumor cells.

In summary, this short but definitive study points to a growing and undeniable evidence in support for T cell based cancer immunotherapy. By incorporating donor derived TCR specificity this strategy vastly expands the reach of T cells immunotherapy. The challenge remains how to streamline this process (exome sequencing, MHC:peptide binding prediction, TCR identification, TCR transduction and re-infusion) to make it affordable for every cancer patient.  

David Usharauli

  

PSGL-1, a ligand for the selectin family of receptors, controls T cell immunopathology

The Selectin receptors, L, E and P, control T cell migration. New study in Immunity showed that Selplg-KO T cells (lacking selectin receptor ligand PSGL-1) display enhanced effector differentiation and greater control of chronic viral infection and tumor, though it comes at the expense of tissue pathology.  

In this paper the authors referred to PSGL-1 as checkpoint inhibitor. However, unlike CTLA4 and PD1 deficient mice, Selplg ^-/- naive mice don't show any abnormality. Interestingly, the authors found that Selplg ^-/- mice efficiently controlled chronic LCMV infection (C13).

However, this enhanced protection against chronic LCMV infection led to severe tissue pathology.

Additionally, and most likely reason why this paper ended up in Immunity, the authors showed that Selplg ^-/- mice could better control tumor (Yumm1.5 melanoma cells).

At present it is not clear whether PSGL-1 signaling works as an independent "checkpoint inhibitor" in vivo or modulates functionality of other canonical checkpoint inhibitors such as PD-1 .

David Usharauli

Experimental malaria vaccine shows 55% efficacy in small trial

This week Nature Medicine published results from small trial involving controlled human malaria infection (CHMI) and experimental malaria vaccine developed by Sanaria (Sanaria Inc., Rockville, Maryland, USA). This vaccine based on attenuated Plasmodium falciparum sporozoite (abbreviated as PfSPZ vaccine) showed 55% efficacy at 1 year post-immunization.

Currently, only malaria vaccine available on market, RTS,S, has efficacy of ~ 22%. In this new study, scientists tested different dosages and routes of immunization for new PfSPZ vaccine as follows:

The results of the controlled human malaria infection are shown below. Here, healthy malaria-naive volunteers were first vaccinated and then received malaria infection from actual mosquitoes bites that carry malaria clone 3D7. The best outcomes were achieved with groups 4 and 5 (i.v. administration, 4 doses of 2.7 × 10⁵ PfSPZ).

Next step was to understand immunological correlates of protection. However, this task was quite challenging, as it turned out. All vaccination protocols, with the exception of i.m. immunization, induced anti-malaria Ab or T cell responses. However, when vaccine trial participants were divided based on blood parasitemia following malaria challenge, it was observed that participants without parasitemia developed higher levels of anti-malaria serum Abs.

At the cellular level, however, the only marker that correlated with vaccine efficacy was frequency of unstimulated Vγ9+Vδ2+ γδ T cells which comprises ~75% of γδ T cells in blood. Actually, the frequency of Vγ9+Vδ2+ γδ T cells in pre-vaccinated individuals was the only marker that correlated with vaccine efficacy in challenge model.

In summary, this study showed that there is still room to improve malaria vaccine. Vaccine efficacy of 55% in 1-year followup is a significant progress when considering that In 2015 there were an estimated 214 million clinical cases of malaria in the world. Another outcome of this study is that fact that it is quite hard to find or define immune correlates of protection and this is especially true for parasitic infection such as malaria or dengue.

David Usharauli

T cell compatible cancer chemotherapy synergizes with cancer adjuvant vaccine

Nowadays it is well acknowledged that optimal cancer treatment would require multi-pronged approach, for example, combining cancer chemotherapy with cancer antigen-specific vaccine to boost tumor-specific T cell response. However, such combination is not always feasible since (a) both tumor and activated T cells are highly proliferative cell types and (b) vast majority of chemotherapy drugs target cellular replication machinery. Basically, it is matter of trial and error to find T cell compatible chemotherapy drugs.

Such combination was recently described in new paper published in Science Translational Medicine. Here, the authors showed that cervical cancer treatment with CarboTaxol (carboplatin + paclitaxel) synergizes with therapeutic T cell vaccine that consisted of HPV16 overlapping synthetic long peptides (HPV16-SLPs) admixed with adjuvant Montanide ISA-51.

Most cervical cancers in humans are induced by human papillomavirus type 16 (HPV16). Initially, the authors showed that CarboTaxol synergized with HPV16-SLP vaccination in HPV16-positive TC-1 tumor-bearing mouse model.

Next, the authors noticed that CarboTaxol [and vaccine too] reduced level of CD11b^highGr1^high myeloid [suppressor] population within tumor tissue.

Similar depletion of myeloid population was observed in blood samples of cervical cancer patients undergoing CarboTaxol therapy.

In fact, CarboTaxol treatment cycles significantly improved T cells proliferation in response to minor [bacterial recall antigen mixture, MRM)] and major HLA antigens [MLR].

Finally, combining CarboTaxol treatment with HPV16-SLP vaccination improved T cell stimulation in response to HPV16 antigens E6/E7.

In summary, this study indicates that CarboTaxol chemotherapy augments, rather than inhibits, tumor-specific T cell priming in response to HPV16-SLP vaccination.

David Usharauli

Spontaneous death of myelin-producing oligodendrocytes could trigger delayed MS-like symptoms

I usually don't check Nature Neuroscience. It does not typically publish immunology-related studies [though MS studies are of interest]. So I was surprised to see such "heavily" packed immunology article there and decided to review it to understand how it ended up there.

It is immediately clear that is not written from immunologist point of view. In general, inter-disciplinary studies are encouraged but if it is not done properly it produces lesser quality research. 

Now, in this paper the authors found that conditional deletion [by death] of brain glial cells, oligodentrocytes, who are responsible for myelin production, triggers late onset MS-like neurological symptoms.

To this end, the authors have used Plp1-Cre-ER^T;ROSA26-eGFP-DTA mouse model where tamoxifen injection releases stop signal from diphtheria toxin A production in oligodendrocytes leading to their death. It appears that this is a rare model of oligodendrocyte deletion where mice actually survive long-term (but it appears this DTA model show late-onset [starting at weeks 26] "leakiness" in absence of tamoxifen injection. This knowledge in itself creates host of issues in data interpretation).  

So, the authors noticed that starting 40 weeks post tamoxifen injection [but not at 10 weeks], spleen and cervical lymph nodes of Plp1-Cre-ER^T;ROSA26-eGFP-DTA mice contained MOG-specific effector T cells. These were accompanied with clinical symptoms of EAE.

Similar results were found with 2D2 transgenic CD4 T cells [specific for MOG] transferred into tamoxifen-treated Plp1-Cre-ER^T;ROSA26-eGFP-DTA host.  

Now, next experiments were quite surprising. To clearly show the role of T cells in the development of late onset MS-like symptoms in tamoxifen-treated Plp1-Cre-ER^T;ROSA26-eGFP-DTA mice the authors tried to cross this DTA model with RAG KO mice. Interestingly, these T/B cell-deficient DTA mice did not survive after tamoxifen injection, implying that recovery from acute oligodendrocyte deletion [following tamoxifen injection] somehow required presence of T or B cells [reminds of studies done by Michal Schwartz lab]. However, the authors neither tried to use CD3KO or B cell KO or simply Ab depletion to test these hypotheses.

Other set of experiments with adoptive transfer of T cells harvested from tamoxifen-treated Plp1-Cre-ER^T;ROSA26-eGFP-DTA mice into RAG deficient mice (but not in WT host) produced MS-like symptoms.

In the remaining experiments the authors tried to show that injection of MOG peptide coupled to nanoparticles could tolerize self-reactive T cells.

So what we learned from this study? First, I am surprised that it even get into Nature Neuroscience [it does not belong there]. Now, this could mean few things: (1) this research was rejected from Nature Immunology and ended up in Nature Neuroscience; (2) It was directly sent to Nature Neuroscience but reviewed by non-immunologists; (3) standards for Nature Neuroscience is much lower compared to Nature Immunology, in general.

David Usharauli

Almost every cancer patient harbors tumor-specific T cells

We know now that adaptive immune system (T and B cells) can detect single amino acid changes in mutated proteins. However, such efficiency of immune system was (and still) in odds with cancer development since tumors invariably express mutated proteins. So what's the deal?

Until recently in vitro detection of tumor-specific immune response was technically challenging. First, detection of mutated proteins or RNA was not easy task. Second, culture conditions for expansion and identification of viable tumor-infiltrated lymphocytes (TIL) specific for tumor antigens were not easy either. Today we have different situation. We can both detect and identify both tumor antigens and tumor antigen-specific TILs.

For example, new paper in Science showed that 9 out of 10 patients with GI tract cancers harbor tumor-specific T cells. However, out of 4 patients receiving adoptive transfer of in vitro expanded tumor-specific T cells, only 1 patient showed persistent anti-cancer response.

This study is kind of follow up from earlier study published in 2014. Here, for each cancer patient, the authors [led by Steven Rosenberg/NIH] first identified cancer mutations by rapid RNA sequencing and designed dozen of tandem minigene constructs containing mutated RNA sequences. These TMGs were then transfected into patient's autologous DCs and co-cultured with multiple TIL cultures harvested from metastatic tumors. This technique revealed presence of tumor-specific T cells (example below, TMG7, 14).

These tumor-specific TCRs were cancer antigen specific [showed no reactivity towards wild-type epitopes].

In 1 patient, the authors found TIL culture reactive to cancer driver gene KRAS (KRAS^G12D). The authors could identify HLA allele presenting mutated peptide (the authors even filed a patent for TCR directed to mutant KRAS. But the TCR itself is a product of nature and not patent eligible and TCR transduction into other T cells is by now considered "prior art". So I am not sure if they have a valid patent claim here. Another point is that it is safe to assume the authors already have earlier patents for transduction techniques and other TCRs. If so, then why they would need a new patent for just another TCR?).

however, the most relevant results are not shown in the paper but just discussed. The authors mentioned that 4 patients were treated with adoptively transferred T cells. However, only 1 patient receiving tumor-specific CD4 T cells showed persistent response and ongoing tumor regression, while other 3 patients who received tumor-specific CD8 T cells either do not show any response or showed only transient response. This suggest that unlike CD4 T cells, tumor-specific CD8 T cells may be not very efficient against solid tumors [lack of persistence in transferred hosts].

In summary, this article supports a notion that cancers do not go unnoticed by immune system. It appears that almost every cancer patient harbor tumor specific T cells that can be harvested, expanded and re-introduced back to patients to target tumors [alone or in combination of checkpoint inhibitors]. However, understanding biological difference between effectiveness of tumor-specific CD8 and CD4 T cells surely requires further research.  

David Usharauli