Monthly Archives: April 2014

Silo Busting: Can We Build Better Immunotherapies for the Treatment of Prostate Cancer?

A very interesting paper published last week in Cancer Research. Written by investigators at The Johns Hopkins University Sidney Kimmel Comprehensive Cancer Center, the paper addresses unmet need in prostate cancer (PC), and proposes a multi-modality approach to building an effective therapeutic. The authors are Nathaniel Brennen, Charles Drake and John Isaacs and title is Enhancement of the T-cell Armamentarium as a Cell-based Therapy for Prostate CancerThe abstract is here.

The authors review the state of hormone-treatment and chemotherapy refractory, metastatic, and/or relapsed prostate cancer, collectively mCRPC. Approximately 200,000 patients (male) are diagnosed with PC each year in the US. Deaths from mCRPC number roughly 30,000 men/year in the US, and there are very limited treatment options. Potential therapies include PC vaccines like Provenge and others in clinical development (GVAX, ProstVax).  Notable for anticipating the need for broad immune stimulation ProstVax uses viral vectors which encode prostate-specific antigen (PSA) along with costimulatory molecules B7.1, ICAM-1, and  LFA-3. More conventionally, Provenge and GVAX are composed of prostate cancer cells that are irradiated and then engineered to express the immune molecule GM-CSF. Provenge uses patient-derived tumor cells while GVAX uses PC tumor cell lines. Provenge offers a modest survival advantage versus the standard of care chemotherapeutic, docetaxel. GVAX failed to meet its primary end point of overall survival when compared with docetaxel in a phase III study. In a phase 2 study, ProstVax treatment led to a median overall survival (OS) of 25 months, very similar to Provenge (vs. 17-22 months in the placebo groups, respectively). Many other variations on this methodology are in development. Other immunotherapeutic approaches in the pipeline include antibodies such as Vervoy (ipilimumab) and nivolumumab that have shown efficacy in other solid tumors. The anti-CTLA4 antibody ipilimumab, a potent immune checkpoint modulator, is being evaluated in mCRPC, both alone and in combination with chemotherapy, radiotherapy or tumor vaccines. Current phase 3 trials include ipilimumab vs placebo in chemotherapy-naïve patients and a second trial in patients who have first received chemotherapy and bone irradiation. Early results suggest that previously treated patients obtain minimal benefit from ipilimumab but that newly diagnosed patients perhaps fare better. Surprisingly, early clinical studies with the anti-PD-1 antibody nivolumab failed to show any benefit. Another approach will be to build T cell therapies such as those based on chimeric antigen receptors (CARs) that have been successful in treating leukemia and lymphomas. However, while known mCRPC antigens are certainly overexpressed in tumor cells, they are not uniquely expressed. For example folate hydrolase (PSMA), a targeted mCRPC antigen, is also expressed, albeit at low levels, in the kidney, small intestine and both the central and peripheral nervous systems. The prostate stem cell antigen (PSCA), another mCRPC antigen, is also expressed in the bladder, colon, kidney, and stomach. Given the inherent aggressiveness of CAR T cells, targeting these antigens is dangerous business indeed.

Back to the paper. Instead of just stating that we need to mix novel immunotherapeutics until we find the right combination, the authors spend some time deconstructing this particular tumor type, and then rationally build up a therapeutic hypothesis based on what the tumor has shown them. This is of course the essence of the personalized medicine approach, although perhaps based more on critical thinking about the biology rather than cataloging driver mutations (both being needed in the bigger picture). And while their ultimate proposals may be overly engineered, they are certainly on an interesting track.

So what do PC tumors tell us? First, PC is often an indolent and slow growing tumor that is caught early by routine screening for soluble prostate specific antigen (PSA, we’ll come back to all these antigens later). Second, more than 80% of PC tumor biopsies have extensive inflammatory infiltration, showing that the immune system has recognized that something is wrong, but can’t seem to fix it. Third, the majority of cells within the inflammatory infiltrate are T cells, so we have the right cell population sitting there ready to kick off a defense (but they don’t).

This is a situation that resembles that of melanoma, that is, a solid tumor that is filled with disabled immune cells, many of them T cells. The difference is that melanoma can be effectively treated with ipilimumab, nivolumab and, very strikingly, with the combination of the two. We reviewed the synergy of immunotherapy treatments for melanoma in our prior post.

So why are PC tumors different? The T cells appear either “exhausted” and unresponsive (anergic T cells), or are actively immuno-suppressive T-regulatory T cells (Tregs). They are identified by expression of specific transcriptional factors (FoxP3) and cell surface proteins (CTLA4, PD-1, and others). They respond preferentially to specific factors that keep them quiescent or immuno-suppressive (IDO, TGFbeta, IL-10). Basically they are so loaded down with redundant and compensatory off-signals that they simply will not be roused by any single method – not a vaccine, not an immune-checkpoint antibody, not even, it seems, when given in combination. A bigger stimulus is needed.

And this is where Brennan et al. lead us in their recent paper. They begin by introducing a CAR built by the team at Memorial Sloan Kettering Cancer Center in New York, led by Michel Sadelain. As a reminder, most CAR constructs contain a single domain for antigen recognition, usually derived from an antibody. We discussed the technology earlier (see this post, and related posts from March). So, for example, the CAR constructs used to transduce T cells that successfully treat acute and chronic leukemias (ALL and CLL) all recognize the antigen CD19. The CAR proposed by the MSKCC group has two antigen recognition sites, one for PSMA and one for PSCA. As noted above, neither of these antigens alone is specific for mCRPC, nor would I imagine the combination to be 100% specific. As someone tweeted during AACR: “A PSMA CAR? Their gonna kill someone with that drug”. Suffice to say, that’s one problem with the dual-targeted CAR. Another, more theoretical, is presented by Brennen et al. as a problem of effectiveness. This is a curious argument. The author’s state that all CAR therapies “share a dependence on endogenous T-cell effector functions”. This is maybe an unfortunate choice of “endogenous” since the activity of CAR-T cells is certainly not endogenous but rather artificially driven by the construct with which it is transduced. The available data suggest that such activity is considerable and dependent only on the availability of antigen. Whether CARs will shut down once they encounter the immunosuppressive environment of the mCRPC tumor is a different question that has nothing really to do with endogenous effector function. But lets move on and take their point for the sake of discussion as it drives the next interaction of their proposed therapeutic, which is to introduce a non-natural cytotoxic element to the CAR.

I suspect the authors are being provocative to make a point, but this seems a bit overdone. Still, lets soldier on. The proposal is now to create a Trojan Horse out of a CAR T cell, by arming it with a proform of a toxin. The example offered in proaerolysin, a vicious bacterial protein capable of blowing holes in every cell it encounters. The toxin is known from studies in which it was injected directly into tumors, in which setting it destroys every cell in a very small area, thus ineffectively. A second iteration of this strategy was to mutagenize the toxin by introducing a PSA-dependent cleavage site, thereby only releasing the active toxin in the presence of PSA, the PC selective biomarker used in routine testing. This Trojan Horse or “molecular grenade” creates a “kill zone” when cleaved by PSA.

To recap. We now have a dual CAR targeting PSMA and PSCA, carrying proaerolysin, that is activated by PSA. It actually get more complex then that, but lets just stop there, take a step back from this Rube Goldberg approach, and ask if we can find merit in the exercise.

There are two interesting concepts here, useful in their own right. One is to “mask” the CAR, irrespective of Molecular Trojan Kill Zones. One genuinely scary and unpredictable aspect of CAR technology is off-target toxicity. This has been seen repeatedly, and casts a long shodow over every new CAR antigen in development. Why? Because there is the “what if” aspect. What if the antigen is expressed somewhere else, unexpectedly, or what if the domain used for antigen recognition also sees a different, related, antigen on normal tissue? Both of these things have happened, and when these things happen, patients die, very quickly.

So, masking a CAR, even a single domain CAR is a very good idea. Happily this is a well worked field, with companies like CytomX busy working out the utility of using tumor-specific proteases to cleave specific peptide sequences that otherwise effectively mask antibody-drug complexes (ADCs) in an effort to improve their safety. That technology may be transferable in some form to CAR technology, providing an added layer of specificity without having to add up all these different tumor-selective antigens (PSMA, PSCA, PSA).

The second is to “arm” the CAR, masked or otherwise, with a toxin. This takes some thought but is possibly an attractive idea, if one could work out the geography (where to place the thing so it works where one wants it to work). I’m a little puzzled about the idea that you could add a toxin, in proform or otherwise, and avoid killing the transduced T cell. For example, PSA, used in the thought experiment above, exists in systemic circulation, a bad place to blow up your Trojan Horse.

Regardless the take-home message is that we might productively combine technologies across the immunotherapeutic space, to build better individual therapeutics.

Returning to mCRPC, where does immunotherapy actually stand, today. It’s fair although sad to say that progress here is similar to most other solid tumors, that is, there has been limited success. There is a vast amount of work to be done but happily there is an army of researchers and physicians willing to do it. With any luck at all papers like the one discussed in this post will help point the way, as loopy as a Molecular Trojan Kill Zone might sound today.

We’ll talk about iCARs next perhaps. or maybe BiKES. We’ll see.

stay tuned.

Novel Synergies Arising in the Immunotherapy of Melanoma

Steven Rosenberg gave an interesting talk at this year’s American Association for Cancer Research meeting (AACR 2014). He discussed various cell therapies that were developed at the National Cancer Institute (NCI). He began with a review of 3 trials in metastatic melanoma that used the patient’s own tumor infiltrating lymphocytes (TILs), isolated, expanded and re-injected, as the treatment. Ninety-three patients were enrolled in the trials. The partial response rate (PR) was 32% and the complete response rate (CR) was 22%. Notably, some of the CRs were durable; Dr Rosenberg went so far as to state that TIL therapy could be curative, albeit in a relatively low percentage of patients treated. In a new trial of 110 patients they are seeing similar results, including durable PRs.

Similar attempts to use TIL therapy in other solid tumors have mainly failed. So one interesting question, posed by Dr Rosenberg, is why do melanomas readily respond immune therapies? Such therapies include not just TIL-based treatment but also to high-dose IL-2, checkpoint inhibitors: blocking CTLA4, blocking the PD-1 pathway, even agonist anti-CD40 antibody (mAb) treatment. All of these therapies will activate cytotoxic T cells and should also activate the rest of the immune system either secondarily, or in the example of agonist anti-CD40 mAb therapy, directly.

Melanomas are unusual in the abundance of TILs that are found within the tumor and the tumor microenvironment. Rosenberg floated the “mutation” hypothesis to explain why TILs are abundant in melanoma: melanoma tumors are highly mutated, with an average of 34 mutations per individual patient tumor. The mutation hypothesis posits that it is the abundance of mutations and therefore mutated proteins that drive TIL accumulation, that is, the mutations produce antigenic protein fragments that can be presented in context of MHC (MHC class I and class II are complexes found on antigen-presenting cells that activate T cells).

If this hypothesis is correct than several predictions can be made. One is that we should be able to find antigenic peptides that activate the TILs from specific patients. Another is that the TILs should be disabled by the tumor or tumor microenvironment (this is already suggested by the success of immune checkpoint inhibitors like ipilimumab and nivolumab in melanoma). Indeed, TILs isolated from patient melanomas express multiple immune control pathways, both in the immune response inhibitory pathways (PD-1, CTLA4, TIM-3) but also immune response activation pathways (4-1BB, OX-40, CD25, CD28, CD27, CD70) and others (LAG-3). So, these calls appears primed to respond, but are held in check.

Further, the TILs are primed to respond, at least in part, to tumor-derived peptides. Dr Rosenberg and colleagues sequenced the tumors from individual patients and used an algorithm to scan the data and identify immunogenic peptide fragments. They then synthesized the peptides and ask whether any of them could stimulate patient TILs. For each patient they found several immunogenic peptides. They could then isolate the T cell receptor (TCR) that mediated that recognition, and use it in an expression construct to develop mutation specific T cells. Note here that it is the TCR on the T cell that interacts with the MHC complex on antigen-presenting cells to trigger T cell activation. We have moved now from bulk TILs expanded ex vivo and re-injected to patient-specific engineered T cells specific for tumor antigens. This TCR-based cell therapy has now shown activity beyond melanoma and may be useful for other solid tumors that contain large populations of TILs. Finally, it may also be feasible to use the TIL immunogenic peptide data to craft highly tumor specific CAR constructs, i.e. by raising the CAR Vh domain (engineered as a scFV) to tumor-mutated antigens.

There remain significant unanswered questions. Other tumor types carry very high mutational burdens but do not accumulate large numbers of TILs – why not? The expression of immune control pathways on TILs derived from melanomas is complex – how best to manipulate these pathways? Also, how do TIL immune control phenotypes vary among patients? The identification of patient-specific immunogenic peptides may be useful in moving tumor vaccine therapy forward – how best to incorporate this data? Finally, a theme we always return to – how should doctors and patients use TCR-based therapeutics in the context of other available therapies.

The TIL data remind us that tumors raise an immune response to tumors, and this has implications for the re-emerging tumor vaccine field. Perhaps these mutated tumor antigens could be used in the context of tumor vaccination. There were several talks at AACR14 describing successful application of tumor vaccines in early phase clinical trials. There have been high-profile failures in this space – GSK’s phase 3 bust with their MAGE-A3 vaccine being a notable recent example. But sticking to melanoma, we see a few strong signals emerging.

Roger Perlmutter updated results from Amgen’s Phase 3 trial with T-Vec, which was initiated during his tenure (he is now at Merck). The T-Vec program was brought into Amgen with the $1 billion buyout of BioVex. T-Vec is a engineered viral vaccine that can infect and then replicate in tumor cells, pumping out the pleiotropic, immune-system priming growth factor GM-CSF along with encoded antigen. The injection is given at accessible tumor sites, e.g. in the skin, causing the melanoma to shrink. Importantly, not just the injected tumors, but tumors distant from the injection site responded, indicating that a systemic immune response had been triggered. T-Vec was compared to GM-CSF injection alone. While the overall response rate was high (about 60%) the interesting data are the comparisons of duration of response.


time to progression or death (primary endpoint)

       overall survival (OS)         (a secondary endpoint)


2.9 months

19 months


9.2 months

23.3 months

The response can be traced to cytotoxic T cells. These initially resemble patient TILs. However, after immunization these T cells have up-regulated immune response proteins (CD28, CD137, CD27, GITR) and down-regulated immune checkpoint proteins (PD-1, CTLA4, Lag3, TIM-3). So this immunization protocol is resetting the T cell phenotype, from immunosuppressed or anergic, to immune-competent and activated. This biological response is likely driven by the effect of GM-CSF on monocytes, macrophages and related cells. The mechanism of action bears further study.

We have not seen enough data yet to determine if there will be long-term responders (those that contribute to the “long tail” phenomena on OS curves) as we see in the immune checkpoint inhibitor trials. Regardless, Amgen is moving forward with clinical trials of T-Vec in combination with anti-CTLA4 mAb (Vervoytm, from Bristol-Myers Squibb) and with anti-PD-1 mAb MK-3475, in collaboration with Merck.

Lindy Durrant and colleagues from the University of Nottingham used a different approach to engage the immune system in the vaccine setting. They developed SCIB1, a DNA immunotherapy that encodes epitopes from gp100 and TRP-2 (melanoma antigens) into a human IgG1 antibody (honestly I need to understand better how they engineered this). The DNA vaccine is electroporated directly into muscle weekly x 3 and then at 3 months and 6 months. The transfection results in expression of the construct that is then taken up by Fc-receptor bearing cells via the CD64 Fc-receptor. CD64+ cells include monocytes, macrophages, dendritic cells and other immune cells. This Phase 1 study was designed as a 3×3 dose escalation study with an expansion cohort at the maximum tolerated dose, determined to be 4mg. Stage III and Stage IV melanoma patients were enrolled. 19/20 patients were shown to have an immune response to vaccination. There was a clear dose response. In the expansion cohort (n = 14) all patients showed an immune response despite expression of PD-L1 on tumor cells. Epitope recognition by both CD4 and CD8+ T cells was observed. Median survival of the expansion cohort is currently 15 months.

While this is a small early stage trial, such results are dramatic and highlight the concept that productively engaging the immune response requires recruitment of the patient’s antigen presenting cell populations (as noted above in the T-Vec example, this is what GM-CSF does). The tumor cell profile data hint at the potential use of PD-1 pathway blockade as a co-therapy for this DNA vaccine approach.

For smaller companies developing cancer vaccine modalities the potential to develop their technology alongside immunotherapy agents should be attractive. While PD-1 and CTLA4 targeting antibodies remain one obvious approach, data presented at AACR suggest that immune activating pathways (GITR, OX40 and others) might also be useful in the context of immune vaccine approaches. The trick will be to aim carefully.

We’ll follow up with a look at immune activation pathways.

stay tuned.

Three high-altitude take aways from AACR14

The American Association for Cancer Research (AACR) 2014 meeting last week was high energy and high impact. We will dive into particular talks and specific pathways and indications in later posts, in the meantime I wanted to mention a few key themes.

1) Immunotherapy Versus The World.  That’s a deliberate overstatement of a subtle shift in emphasis from last year’s big meetings, where combinations of immunotherapy with just about anything else were the hot topic. This year there were several talks which emphasized the futility of chasing oncogenic pathways and all of their resistance mutations, one after the other, as opposed to letting the immune system do the work. However, it seems to me overly optimistic to believe that immune modulation can defeat a high percentage of patient  tumors on its own, as some speakers acknowledged. Combinations remain necessary although we will have to work past some notable failures in combo trials, such as the liver toxicity seen in the ipilimumab + vemurafenib combination phase 1, discussed briefly by Antonio Ribas               (see

2) Immunotherapy Versus Itself.  In the ultimate battle of the titans, we see different immunotherapeutic modalities squaring off. This is a theme we’ve touched on before in this space, but the  competition is getting heated. In some indications, the leukemias, lymphomas, perhaps melanoma and some other solid tumors, there is an abundance of therapeutic choices, and the hard question of which therapy best suits which patient will ultimately need to be addressed outside of the context of clinical trial enrollment. Several talks really brought this message home. Roger Perlmutter of Merck (and before that, Amgen) envisions an important role for multiple immune therapies including bi-specific antibodies, chimeric antigen receptors (CARs), and immune checkpoint modulators like Merck’s anti-PD-1 antibody MK-3475.  For B cell lymphoma for example, there is blintumumab (Amgen), a potent bi-specific that redirects T cells to CD19+ tumor cells (and normal B cells), and there is CTL019, a CAR therapeutic which does much the same thing. The therapeutic profiles and toxicity differ, but the general idea is the same. One big difference is that while CTL019 drives T cell expansion and the development of long term anti-tumor memory, the bi-specific does not. Which is better? We don’t know yet. He did not mention that one might do well trying a course of BTK inhibition plus anti-CD20 antibody therapy, perhaps with restricted chemotherapy first e.g ibrutinib plus rituximab and chemo (R-BR or R-F). That choice comes down to efficacy, then toxicity, and eventually cost. Efficacy seems to be a home run with the CAR therapeutics, although these may run into trouble in the area of toxicity and cost calculation. Renier Brentjens discussed the CAR therapies being developed under the Juno Therapeutics umbrella. Acute lymphoid leukemia (ALL) can be treated with CAR 19-28z modified T cells to achieve a >80% complete response rate with >70% of patients showing no minimal residual disease, an outstanding result. However, 30% of treated patients end up in the ICU due to cytokine release syndrome and other toxicity, and recently patients in the ALL trials have died from unanticipated tox causes. Juno stopped 5 trials of their CAR technology last week due to toxicity. Apparently one patient died of cardiovascular complications and another of CNS complications (severe uncontrolled seizures) – it was hard to nail down as Dr Brentjens had gone off his prepared talk for these remarks which were off the cuff, so comment please if you have better info on this. Carl June discussed Dr Brentjens’ presentation, noting that the clinical results were really quite striking, and contrasting the CD28 motif-based CARs with the 4-1BB-based CARs (as designed by Dr June with U Penn and licensed to Novartis). He also stressed that in chronic lymphocytic leukemia (CLL) they have had patients who have failed up to 10 prior therapies, including rituximab and/or ibrutinib, and these patients have responded to CAR treatment. That’s very impressive data. The roadblocks to widespread use of CAR therapy however are large and include the toxicity, the “boutique” nature of the current protocols, the cost. Perhaps, Dr June suggested, CAR will end up as third line therapy, reserved for salvage therapy. I for one hope not.

Also in the immunotherapy space were hot new targets (e.g. CD47, OX40, GITR), advances on the vaccine front, and a few surprises. We’ll update soon.

3) The Medicinal Chemists Have Been Busy.  Not to be drowned out by the Immunotherapy tidal wave, small molecule therapies targeting specific oncogenic pathways continue to be developed and show promise. Most readers will be aware of the high stakes showdown (so billed) between Novartis, Pfizer and Lilly in the field of specific CDK4/6 inhibitors – in addition to bringing forward some really nice phase 2 data (we’ll discuss these another time) this “showdown” also illustrates that current portfolio strategy drives a lot of overlapping effort by different companies. As expected, much of the action is moving downstream in the signaling pathways, so we saw some data on MEK1 inhibitors and ERK1/2 inhibition. There were some new BTK inhibitors, nice advances in the epigenetics space, and some novel PI3K inhibitors. All grist for the mill.

stay tuned.

Update from AACR14: Clinical Halt for Memorial Sloan Kettering/Juno Therapy in Non-Hodgkin Lymphoma

Yesterday we learned that the Memorial Sloan Kettering Cancer Center (MSKCC) and corporate partner Juno had stopped enrolling patients into 5 clinical trials of their chimeric antigen receptor (CAR) T cell therapies. Details are spare at this point, but unexpectedly, the cause of the clinical stop was severe cytokine release syndrome (CRS). I say ‘unexpectedly’ because it was just last month that MSKCC released an update on their ability to detect CRS early enough to initiate aggressive treatment. We commented on this update in a recent post on the CAR 19-28z technology.

According to the MSKCC update given in February, they had developed “guidelines for managing the side effects of cell therapy” including CRS, and “diagnostic criteria” for identifying at-risk patients using clinical lab tests. These tests were for a panel of cytokines and for C-reactive protein (CRP). To be fair these comments were made in reference to work ongoing in acute lymphocytic leukemia (ALL), but it was clear that the clinicians felt they were broadly applicable. It seems now that these comments were premature.

This is a critical issue in the CAR technology field, potentially holding back not just MSKCC/Juno but similar work from U Penn/Novartis and NCI and partners working with Kite Pharma. The syndrome characterized as CRS is a consequence of the massive immune response to the tumor, which is a designed consequence of the CAR technology. CAR-modified T cells are potent cytotoxic agents, and are designed to recruit unmodified T cells to the cause (the so-called bystander effect). This result is the triggering of the acute phase response, and then an outpouring of cytotoxic compounds, pro-inflammatory cytokines, and effector proteins. When allowed to proceed unchecked, the response begins to engulf normal cells and tissues, causing additional cell death, organ damage, and in the most severe cases, death.

The reality is that clinical responses leading to CRS seem to have caught MSKCC/Juno flat footed in at least one clinical trial of Non-Hodgkin’s Lymphoma (NHL) – we note here that stopping 5 trials does not mean that CRS was seen in all 5, but they are related by clinical indication, so it is an obvious precautionary step to take.

What will happen next we cannot know yet, as we have not yet heard the necessary detail. At the very least the MSKCC/Juno NHL programs are in for careful scrutiny. This will impact the clinical development of the technology and slow access for patients. The patients affected are those who are the most in need, nonetheless, the caution is warranted. More broadly, this unexpected turn of events may encourage us to look again at more established therapeutics for NHL, including small targeted molecule drugs, cytotoxic antibodies, antibody-drug conjugates (ADCs) and bispecifics, and of course, combinations of those therapies with one another or with current chemotherapeutics.

And this just in: In the immune checkpoint space we have just learned this morning of the potential for unexpected immune toxicity after long term treatment. Thankfully this appears to be very rare but this too will bear watching.

Our TIM-1 paper on T cell trafficking has published in Immunity

Sugarcone Biotech is pleased to congratulate our collaborators Gabriela Constantin and colleagues on the publication in Immunity of the paper entitled

TIM-1 Glycoprotein Binds the Adhesion Receptor P-Selectin and Mediates T Cell Trafficking during Inflammation and Autoimmunity by Stefano Angiari, Tiziano Donnarumma, Barbara Rossi, Silvia Dusi, Enrica Pietronigro, Elena Zenaro, Vittorina Della Bianca, Lara Toffali, Gennj Piacentino, Simona Budui, Paul RennertSheng XiaoCarlo Laudanna, Jose M. CasasnovasVijay K. Kuchroo, and Gabriela Constantin.

published online ahead of print today, April 3 2014 DOI:

The paper makes extensive use of antibodies and proteins developed by SugarCone founder Paul Rennert, and Biogen Idec colleagues. This elegant work details a previously unknown function of TIM-1 in regulating T cell movement in the inflamed vasculature, thereby controlling local inflammation. This new biology complements the role previously described for TIM-1 in mediating lung allergic responses. Alongside our recently published work in Ebola virus cellular infection, we begin to appreciate the role of TIM-1 in diverse aspects of infection, immunity, chronic inflammation and autoimmunity. See for more on this important protein.

Here is the summary of the paper from Immunity:

Figure thumbnail fx1



  • •TIM-1 mediates Th1 and Th17 cell capture and rolling on P-selectin in vitro
  • •TIM-1 is a major P-selectin ligand controlling T cell adhesion in inflamed vessels
  • •Both mucin and IgV domains of TIM-1 are required for the interaction with P-selectin
  • •TIM-1-mediated adhesion controls autoimmune and inflammatory disease development



Selectins play a central role in leukocyte trafficking by mediating tethering and rolling on vascular surfaces. Here we have reported that T cell immunoglobulin and mucin domain 1 (TIM-1) is a P-selectin ligand. We have shown that human and murine TIM-1 binds to P-selectin, and that TIM-1 mediates tethering and rolling of T helper 1 (Th1) and Th17, but not Th2 and regulatory T cells on P-selectin. Th1 and Th17 cells lacking the TIM-1 mucin domain showed reduced rolling in thrombin-activated mesenteric venules and inflamed brain microcirculation. Inhibition of TIM-1 had no effect on naive T cell homing, but it reduced T cell recruitment in a skin hypersensitivity model and blocked experimental autoimmune encephalomyelitis. Uniquely, the TIM-1 immunoglobulin variable domain was also required for P-selectin binding. Our data demonstrate that TIM-1 is a major P-selectin ligand with a specialized role in T cell trafficking during inflammatory responses and the induction of autoimmune disease.


EBOLA virus infection paper published in Journal of Virology

In collaboration with Wendy Maury’s lab at the University of Iowa we have published a follow-up paper describing our work characterizing the functional domains of TIM-1 required for infection of cells by the Ebola and Marburg filoviruses. This work may one day lead to the development of therapeautics to stop or control Ebola outbreaks, such as the one occurring now in the West African country of Guinea. Congratulations to Sven Moller-Tank, Lorraine Albritton (University of Tenneesee) and Wendy Maury on this important work.

here is the link:

here is the abstract:

Characterizing functional domains for TIM-mediated enveloped virus entry. 

Sven Moller-TankLorraine M. AlbrittonPaul D. Rennert and Wendy Maury* Department of Microbiology, University of Iowa, Iowa City, Iowa, USA, Department of Microbiology, Immunology & Biochemistry, University of Tennessee Health Science Center, Memphis, Tennessee, US,  SugarCone Biotech LLC, Holliston, Massachusetts, USA. Journal of Virology 2014. Published ahead of print 2 April 2014, doi: 10.1128/JVI.00300-14


T-cell immunoglobulin and mucin domain 1 (TIM-1) and other TIM family members were recently identified as phosphatidylserine (PtdSer)-mediated virus entry enhancing receptors (PVEERs). These proteins enhance entry of Ebola virus (EBOV) and other viruses by binding PtdSer on the viral envelope, concentrating virus on the cell surface, and promoting subsequent internalization. The PtdSer binding activity of the IgV domain is essential for both virus binding and internalization by TIM-1. However, TIM-3, whose IgV domain also binds PtdSer, does not effectively enhance virus entry indicating that other domains of TIM proteins are functionally important. Here, we investigate the domains supporting enhancement of enveloped virus entry, thereby defining the features necessary for a functional PVEER. Using a variety of chimeras and deletion mutants, we found that in addition to a functional PtdSer binding domain PVEERs require a stalk domain of sufficient length, containing sequences that promote an extended structure. Neither the cytoplasmic nor transmembrane domain of TIM-1 is essential for enhancing virus entry, provided the protein is still plasma membrane bound. Based on these defined characteristics, we generated a mimic lacking TIM sequences and composed of Annexin V, the mucin like domain of α-dystroglycan, and a glycophosphatidylinositol anchor that functioned as a PVEER to enhance transduction of virions displaying Ebola, Chikungunya, Ross River, or Sindbis virus glycoproteins. This identification of the key features necessary for PtdSer-mediated enhancement of virus entry provides a basis for more effective recognition of unknown PVEERs.

IMPORTANCE (nontechnical, 150 word limit): T-cell immunoglobulin and mucin domain 1 (TIM-1) and other TIM family members are recently identified phosphatidylserine (PtdSer)-mediated virus entry enhancing receptors (PVEERs). These proteins enhance virus entry by binding the phospholipid, PtdSer, present on the viral membrane. While it is known that the PtdSer binding is essential for the PVEER function of TIM-1, TIM-3 shares this binding activity but does not enhance virus entry. No comprehensive studies have been done to characterize the other domains of TIM-1. In this study, using a variety of chimeric proteins and deletion mutants, we define the features necessary for a functional PVEER. With these features in mind, we generated a TIM-1 mimic using functionally similar domains from other proteins. This mimic, like TIM-1, effectively enhanced transduction. These studies provide insight into the key features necessary for PVEERs and will allow for more effective identification of unknown PVEERs.