This transcript is automatically generated.

Manju George 0:00
Hello everyone. Welcome to DocTalks. I’m Manju George, the scientific director of COLONTOWN. Today we have with us Dr Ingrid Lou and Dr Tim Starr. Dr Lu is an associate professor of medicine at the Division of hematology, oncology and transplantation. He completed medical school at the State University of New York, and finished his residency from Duke University Medical Center, and he finished oncology and hematology fellowship from Memorial Sloan Kettering, and then a neuro oncology fellowship from Duke University Medical Center. Dr Lou welcome, and we also have with us. Dr Tim Starr. He’s an associate professor in the Department of Obstetrics, Gynecology and women’s health. I was looking at his research, and looks like he’s interested in understanding the genetic basis of cancer in order to develop individualized, targeted therapies. So thank you both for being with us, and welcome Dr Starr

Dr. Starr 1:10
I know everybody here is personally associated with colon cancer. So what we’re going to hear talk about today is a clinical trial that’s led by Dr Lu to treat advanced stage colon cancer, and I’m going to give you the kind of basic science behind the trial. Now you’ll have to bear with me. I am a ex high school teacher, and I’ve also taught a lot of undergraduate and graduates, and I’ve learned from that to start at the very basics and never assume anybody knows anything. So for those of you that know a lot more, bare with it. So first off, the basics of this trial is it’s an adoptive cell therapy trial. And the key here is the therapy is cells, not drugs. And so the basic backbone of an adopted cell therapy is you take cells, and you can take them from the patient or possibly from a donor. In this case, we’re taking them directly from the patient. And for adoptive cell therapy, it can be any types of cells. It could be a T cell. Could be B cells, it could be natural killer cells, stem cells. What we’re focusing on in this trial, we’re calling tumor infiltrating lymphocytes, really just a mish mash of immune cells that are hiding out in the tumor. And then you take the cells out, you do something to make them better at killing the tumor. And this can involve lots of different things. People are trying lots of ways. You could just make more of them, expand them, enrage them somehow, by giving them cytokines, select out the ones you think that are going to do the best killing you can then now add genes and delete genes. We have the technology, which I’m going to talk about a little bit later, and then you treat the patient with those enhanced cells. So that’s the nutshell of adoptive cell therapy, and that’s what this trial is. Now it’s immune cell therapy. So I have to give you the basics. 101, on immune therapy, which I hear you’ve already seen some people give talks on this. So just remember that your body has T cells. T cells have this receptor on their surface. Think of it as a mutation detector. It’s also a virus detector. Turns out that, you know, 1t cell is going to detect one type of mutation or one type of virus. Luckily, you have 1000s to 10s of 1000s to millions of these, each one with a different flavor of a receptor. Now you probably hear the trial in the background there, it’s Wednesday at 1:00. So the way we’ve evolved is that healthy cells actually show their guts to these T cells that are patrolling your body. And the way they do that is they kind of chop up the proteins inside them and put them on these special molecules that display them out to the T cells. Now your T cells have been trained since birth not to recognize healthy cell guts, so any proteins that have been around shouldn’t have a T cell that’s going to try and react with it. So the key here is, with tumor cells, some of these genes get mutated. You know about this, and so the guts are now going to have some mutated proteins, and that mutated protein will get shown and instead of calling it an epitope, now we’re going to call it a NEO epitope, or a NEO antigen. And if the immune system is working, hopefully there’ll be a T cell somewhere that has the matched mutation detector for that mutation, and it will recognize that tumor cell and kill it. Of course, if the tumor is around, it’s probably not working all that well. This is kind of the not so basic so, so when that T cell does recognize that tumor, how does it actually go and kill it? So it turns out there’s a huge slew of proteins inside T cells, signaling pathways that need to be activated, some need to be turned off, for the T cell to actually physically go and try and kill that tumor cell. And it can do that by creating little holes in the tumor cell or throwing molecules at it that kind of tear it apart, apoptosis. So lots of signaling going on. So the basics for our trial is we want to use tumor infiltrating lymphocytes put back in the patient to kill the tumor. But the first thing we want to do is, we want to select out those tumor infiltrating lymphocytes that we think are going to attack the tumor. So here’s the schematic for the trial. We’ll biopsy the patient, so we get a chunk of tumor. The tumor should have these infiltrating lymphocytes in it. Now they’re not working, because this patient has a tumor that’s growing wildly. So they’re there, but they’re not doing so well. So what we do is we cut the tumor into chunks, and we actually grow a little bit of the tumor in its own private well. So we have these 48 well plates, so now that tumor infiltrating lymphocytes that were with that chunk will grow in that well and so forth. And we help them along a little. We encourage all these lymphocytes to grow by giving them a candy they like, called interleukin two, IL2, which stimulates their growth. So we get these wells with these pills growing. Now what we can do is we can actually sprinkle them with the Neo epitopes in the tumors and test to see if they can actually recognize those new epitopes. So I skipped something here. We also take a little piece of that tumor and we sequence it. We sequence it to find all the mutations that are specific to that one patient, and we physically sprinkle those on to the tills. Now this is showing you one plate with six wells. We’re actually doing this in about 100 plates. Each plate has about 96 wells. So it’s a big test. Takes us a little while to do this. But what we do is we find the wells that seem to react to those new epitopes, and then we can pull out those tills, expand them up and throw them back into the patient. And this is kind of what’s been going on for a little while. This was kind of pioneered at the National Cancer Institute by this guy, Steve Rosenberg, so what are we doing to make this even better? So our hope is we can make these TILs better killers. And I date myself here with The $6 million Man Superwoman, and then Dr Lou is a fanatic of The Avengers, so I had to put in an Avenger there for you. Can we? Can we soup these guys up, make them real killers? So here’s the – not going to be too much scientific data. – This is about it, two figures. So our group and others have found this protein, and it’s called sis, stands for inhibitor of cytokine signaling. It’s a protein that actually blocks T cell killing. And here’s the piece of data that kind of proves this to us, is we do these tests in cells, or we do them in mice. So this is a mouse experiment where we give the mice melanoma, and we have a T cell that actually recognizes melanoma, so we can treat the mice with this adoptive cell therapy, but the T cell that recognizes it doesn’t do a very good job. So this is the purple, purple triangles. So this is the size of the tumor in the mouse, and this is the days after we gave them the cell therapy. And you can see the tumor is growing, and pretty soon it grows so big that it kills the mouse. And we have these controls. So these are the normal TILs that have the wild type sis in them. With another set of cells, we can actually knock out the gene sis. And that’s what the SIS minus minus looks like. And when we put these tills in, the tumor stops growing, and this gives us this is a very aggressive mouse melanoma tumor that’s super hard to cure. So this is really exciting data that says, hey, maybe we should try this in humans, and you do some work to try and figure out what’s actually going on inside the cell. So we know that our protein blocks this signaling pathway here, and when it does that in the T cell, the T cell can’t kill the tumor cell. If we knock out the gene and get rid of it, this signaling pathway is activated, and now the T cell is a much better killer, and it can cause the tumor to go into apoptosis, which is our word for killing the tumor cell.

Dr. Starr 9:38
So this is what we want to do to those tills back here, and so what we do physically is we use this new technology that you might have heard of, called CRISPR cast nine and was the science technology of the year. And basically, you take the cells in a dish, you zap them with electricity while you bathe them with. The CRISPR cas9 components, and the zapping causes the cells to kind of break apart a little CRISPR cas9 step gets inside and does its job. So, what is its job? So, another 101, biology. Here, you’ve got your cell, right? You’ve got the nucleus. In the nucleus, you got the chromosomes with the DNA, and part of that DNA is the gene that codes for this ish protein. So we want to get rid of that using CRISPR cas9 so the way CRISPR cas9 works is you give it this enzyme, it’s this big orange blob here called cas9 and you give it this little string of RNA called the guide RNA, which is the CRISPR part. So you give it the CRISPR and the cas9 and what this little string of RNA does is it has the sequence that knows where the CIS gene is, and so it pulls the cas9 enzyme to that exact spot in the genome, and it creates a break. So here’s the DNA. These are the two DNA strands, and it cuts it, and when that cell tries to repair it, it usually doesn’t repair it very well, and it can no longer make the CIS protein. So the last piece of data I’ll show you is what we want to look at. When we take these cells out of the patient, and we run it through that whole process, and we cut them out. We have ways of looking to see if that sis protein is expressed, and it basically shows up as a black blob on a gel that we run that has protein in it. So these are cells that we didn’t put in the CRISPR cast nine, and you can see there’s lots of the SIS protein. These are cells where we did put this in, and there’s almost none. So this is really good. It’s not easy to do this in T cells that you take from patients, but a group working here at the University of Minnesota, especially Brandon Moriarity and Bo Weber have developed this system to get this to work really well in primary cells, which is kind of the limiting factor, which is why people haven’t been doing this before, also because CRISPR cast nine was just invented a little while ago. So now, if we go back to that diagram I showed you, what we do, we’re going to put in this last step before we put the tills back into the patient. So we’re going to take them, we’re going to CRISPR, cast nine, delete out, sis, get the deleted cells, and those are the ones we’re going to infuse back into the patient. So that’s the basic 101. Of the science behind this experiment. And maybe before I pass it over to Dr Lou, if people have any specific questions about this or more technical details they want explained, I’d be happy to do that.

Participant Question 12:48
That does the location of the tumor come into play, as far as delivering

Dr. Starr 12:56
good question. So we when we take the tumor biopsy, we can actually take it from anywhere. So if the patient has a liver metastasis, a lung metastasis, we’ll go in and take it from there, because that’s easier to biopsy. So we haven’t done this obviously, in enough people to know whether it’s going to affect how how many T cells you get out and whether they’re good T cells or bad T cells, but that’s one of the questions we want to ask. So in one of our patients, we’ve done it from a liver biopsy, and one patient, we’ve done it from a lung biopsy, but we could also do it directly from the colon tumor biopsy too, if we needed to. Is that the question?

Participant Question 13:38
Yeah, and the peritoneum is what I’m thinking of, is if you’ve metastasized into the peritoneum, I know everything’s a little more squishy and not so defined in there, and whether that would complicate things, yeah,

Dr. Starr 13:51
What we really want is a, I’m going to call it a healthy piece of tumor. We want a piece of tumor that’s growing well, because that’s where the T cells will have infiltrated into and we want a fairly big one so we can cut it up into many pieces, so we can get a lot of them. So it really depends on this is going to be very end of all individualized for each patient, but we could take it from the peritoneum too.

Dr. Starr 14:13
Thank you.

Manju George 14:14
Thank you very much for that. There are two questions. One is, what is the typical time to grow the tils and then to infuse them. The next question is, are there risks when the TILs are modified that would impact the health of the cells?

Dr. Starr 14:30
Yes, thank you. So first, first question is, we’re trying to get it as fast as possible. But that being said, it takes the time to grow the tills. It takes time to sequence the tumor. It takes time to edit them and pull it out. And right now we’re at about two months, one month to two months we can, we can from the time we take out the biopsy to the time we can infuse them back into the patient, is about that long we can freeze down the cells. So it’s not limited to soon as we get them, we can then put them into the patient, and then the second remind me the second question, what are the risks of this modification to the health of the cells? That’s always an unknown, and that’s that’s the big worry, obviously, for FDA approval and the cells like this have been put into patients, and one nice thing about these cells is they don’t tend to cause a lot of of reaction, like these car T cells that you’ve probably heard of. But this whole idea of cutting out the CIS gene and seeing what happens is really an unknown, and that is a worry of ours. Of course, we’ve done it in mice, and it doesn’t seem to have affected normal, healthy organs like we would be afraid of, but that’s always a possibility. And you know, this is for metastatic colon cancer, so we wouldn’t do this to healthier patients.

Manju George 15:59
The next question is, can you harvest the pills from lymph nodes? Tumor in the lymph nodes,

Dr. Starr 16:05
generally, these tumors, I’ll let probably Emile answer this better. But the tumors don’t, although there might be some tumor cells in the lymph nodes, we really need a good solid junk of tumor. We want a solid piece, and those usually aren’t found in the in the lymph nodes for colon cancer.

Manju George 16:23
Okay, thank you. Great. That seems to be, yeah, go ahead.

Dr. Lou 16:28
No, I was gonna say I’ve known Tim for nearly a decade, and anytime I hear him speak, I learn a lot. And I wish during his teaching phase, he had been my teacher for biology, I think I would be a whole lot smarter. But thank you. I think, I think what’s really also important is what I had in mind and asking Tim to partner with me for this talk. It’s emblematic of the partnership that occurs for this trial. So I’m a physician scientist. I have an MD and PhD degrees. I do basic science research and translation research, and I treat patients in the clinic, whether it’s with standard of care treatment or clinical trials, when those options are available, and we try the best we can. But you know, the ultimate dream that we that has been said for decades, this term bench to bedside, I think that that term, and that the reality of how that plays out has really evolved over time. It’s not as simple as simple as just taking something from the lab to translation into the clinic, but there’s so many logistical hurdles. And is not just safety, but will something work and but this is this is happening, and that’s what this trial is, really the overall strategy. You know, as Tim said, arising for more from more than half a decade of basic science investigation here at the University of Minnesota, in partnership with those esteemed researchers at the National Cancer Institute who had already done a lot of clinical trial research over the decades, and investigation in the lab for adopted cell therapy in general and tills very specifically. And if we think of this as a next generation of that, I think we’re very proud. I think I speak for Tim and really our whole team that really under the proverbial roof of this gigantic campus in Minneapolis, Minnesota, that allow the basic science research that arose out of here, and collaborators, the NCI, we’ve been able to take from that bench so called to the clinic and have this trial open is really, I think, really puts us at the next frontier. And I’m not saying just at the University of Minnesota, but just as a field of gastrointestinal oncology, we’ve seen from afar that our colleagues who treat patients with hematologic malignancies like lymphoma, leukemia, and they’ve had all these fancy immunotherapy and cellular therapies on the horizon, and they’ve been able to treat patients with this type of therapy, and we’ve been very envious, and there have been Things like immune checkpoint inhibitors, which I’m sure you’re everyone in COLONTOWN has heard a lot about for patients with microli and stable disease, which only comprises barely 5% or maybe a little bit more, of all treatment of all patients, but who qualify for colorectal cancer, but what about the other 85 to 95% of patients? We need to do better, and this scientific effort from bench to bedside as an effort to do that. And there will be more other studies. There will be other studies, maybe looking at sis knockout using CRISPR and other types of immune cells, but here looking at tumor infiltrating lymphocytes, this trial brings, brings across several novelties, and I think we’re going to discover things together. And I think at any clinical trial, if we had the answer already, it would no longer be on clinical trial. I’d have the luxury of being able to treat it, treat patients as standard of care. But maybe the answer for this and other new, innovative treatments will be different, and hopefully it will be two years from now, three years from now, five years and 10 years and decades to come. So I’ll go ahead and share my screen if it’s possible, and I will do slideshow view. So consider this the second half of discussion from Dr starr, and I do want to take a moment so dr Manju, George, thank you so much for the opportunity to meet everybody. I follow COLONTOWN from afar online, and I know you and other. Other members have participated in the ACRS, the American Association for Cancer Research Scientist survivor program, which is 20 plus years strong, welcoming advocates to the AC annual meeting to learn about the science. And I think you and everybody in colintown are are not just advocates from afar, but deeply immersed in learning the science, and not just about clinical trials, but the actual science of it. And I think that’s an immense endeavor. So I really am full of admiration, and I’m particularly grateful to have this opportunity during the onset of colorectal cancer awareness month. So March and blue, we wear the blue as a symbol of support for anyone who has been diagnosed with colorectal cancer of any stage going through treatment now, has gone through treatment. And those who have yet to be diagnosed and yet to go through treatment that will benefit from clinical trials like this one and others, so that the trial itself has a long name. And if you go to clinical trials.gov I can show you that page at the end, briefly, if we have time, this is the NCT number. It’s called a phase one two trial in patients with metastatic gastrointestinal epithelial cancers, administering cancer administering tumor infiltrating lymphocytes, in which the gene encoding sis was inactivated using the CRISPR cast nine system. And thanks to Tim’s in depth discussion, you have the context for where that title comes from. There are many, many dozens of people here at the university who make this work, and that’s from within the labs and from the different centers that I’ll talk about briefly, and also throughout the clinics. I have the great pleasure of leading this investigation, which is here at the University of Minnesota, and some of my co pilots, or CO investigators include Dr David McKenna, who is the head of our molecular and cellular therapeutics department, and we’ll talk about that that’s crucial to how we can produce the T cells and expand them for treatment here at the University Jeff Dr Jeff Miller is a renowned bone marrow transplant specialist at cancer researcher Dr starr here, along with when he mentioned Dr Bo Weber and Dr Brandon mayordi, who have been instrumental to developing the CRISPR cast nine system with great efficiency, and specifically looking at sis, and that’s in collaboration with the National Cancer Institute team. So this is a little bit kind of picking up the baton, but it’s in the context that it’s not that no one has ever used tumor infiltrating lymphocytes or tills or adoptive cell therapy in solid tumors before, but as I said, you know, the hematologic malignancies have been little bit more at the forefront, and we’re catching up. But in previous studies at the National Cancer Institute led by Dr Steven Rosenberg and his colleagues, they really blazed a trail for developing protocols for how this can be used, developing safety, better understanding of the toxicities that were observed that were really coming from the high dose lympha depletion, chemotherapy given prior to administration of the or infusion of the tills, and also high dose IL2. So this is the part of translation from what Tim said he described to you and showed you the in picture form that the T cells were fed by the IL two. He called it the candy. And I like that analogy, because il two is a inflammatory stimulant that the cells, the T cells thrive on, and they can replicate and really be robust and last longer. And that’s part of the clinical protocols as developed that have developed over the course of the last decade or two at the NCI, and it’s preceded by chemotherapy that normally would be given by our colleagues who are treating aggressive leukemias or lymphomas. So it’s not the usual standard of fare for patients with colorectal cancer, but in this case, it’s designed to try and prepare the bone marrow to not just receive the T cells allow them to engraft or and ultimately thrive. And so that combination of the lympho depletion, chemotherapy, providing the tills, and then following that with high dose il two is designed to not just build upon the CRISPR engineering part, which is gene editing as Gene scissors. It’s not just the principle of adding to just giving T cells, but in addition knocking out SIS to magnify and amplify its effect. But to even do it further, you can think of those steps so mutation, reactive, kill, act therapy in patients with progressive metastatic gastrointestinal epithelial cancers, NIH has shown promising efficacy signals now, in terms of what type of cancers that means that’s a long, drawn out description, metastatic obviously means having spread far away from where it started. In other words, stage four disease. Gastrointestinal is self explanatory, basically from mouth to anus, but epithelial cancer is really hones in, for example, carcinomas of the esophagus, stomach, pancreatic adenocarcinoma. I think the key there is it’s not going to include pancreatic neuroendocrine tumor that despite anatomic site being in common, it’s really, in many ways, so many, so much of a different biologic cancer. But in terms of the efficacy, what’s been seen in preclinical The reason is to include epithelial, but not neuroendocrine, and continuing along the digestive tract, small intestine, of course, colony, rectum. Um, and so the adenocarcinomas, when they metastasize and refractory to therapy, would make someone eligible for this study. So some of the additional background in the third line setting that data suggests that about 15% of patients with mest act gi associated carcinomas can obtain objective clinical responses when neoantigen selected tills are part of the equation, even just without the benefit of sis modification. And as Tim mentioned, such as an acronym, means fancy form of regulation of T cell receptor signaling to the patients, at least for the patients, and maybe that’s one of the patients that you mentioned earlier in the conversation. Menju to the patients, remain without evidence of disease, no evidence of disease greater than three years after treatment. And I think this is probably one of the crowds I don’t need to preach to and to explain how remarkable that is, and that’s really going from the exception, and we want to make it the rule with more innovative treatments. Pre clinically, CSG knockout has been shown to improve important cancer selling function, cancer killing functions of T cells and mice and humans. And so the next step, of course, is to go from cells in dishes to animal models, and now we’re at human clinical trials. So this is a single center trial here at the University of Minnesota. It’s a combined phase one two study, because we’re looking to answer the questions, many of which I’ve already heard, which you undoubtedly may have, or anyone watching this as recording may have later, really want to not just look and answer the question, Does this new treatment work in individual patients? But we have to prize most and first and foremost. And what keeps me at night up at night, more than any other endeavor I’ve ever undertaken as an oncologist is we are the ward of patients that have entrusted us with their safety, and we have to make sure to do this as safely as possible. So the purpose of the study is to determine the safety of administering these mutation reactive autologous lymphocytes that, in addition to prior studies, has knocked out the CIS gene, and in addition, has used CRISPR cast nine editing to do this. And the patient population, as specified, are patients with metastatic gi epithelial cancers, not when they’re first diagnosed, but further along in treatment, starting with progression on first line therapy or the equivalent. And as the GI oncologist, I added the term maintenance therapy, which it can often be given. For example, after FOLFOX you can achieve stable disease, and just do 5FU to spare further injury from oxaliplatin is just one example. There will be up to five dose levels initially, of this sis, which I reviewed, the acronym for in kale, means knockout. I’m knocking them out using CRISPR cas9 of the tills. And we want to escalate the dose as is as we’re able to do safely. And if we can prove we can do it safely. We can go to higher doses, then we can then establish a dose and concentration of the T cells that are safe enough to proceed for the remainder of the patients, primarily in the phase two version. So instead of bench to bedside, I think the way some of that paradigm has evolved is clinic to lab, back to clinic. And that’s the strategy and the tactic and the approach for this trial, it starts in the clinic, because everything starts in the clinic. Our observations, how we help patients, and when we receive patients who are interested, or if we identify patients who may be eligible for the trial that we can offer to we have a very extensive screening process with my team in the clinical trials office, from point of entry, from the first phone call to welcoming them into clinic, to determining from a long list of eligibility of whether or not there would be good candidates. And so much of that is predicated on safety. As you know many of you, if you’ve investigated clinical trials, you know it’s, it’s nothing to do with personality or where someone lives. It’s, it’s always the underlying strategy is safety, but sometimes it could be laboratory values that need to meet criteria to indicate safety. It could be how well someone is able to, for example, walk and be able to do activities of daily living, such as showering themselves, be able to feed themselves, make basic foods. These are basic measures of safety going in to this kind of process. So when we identify patients and they meet eligibility criteria, we can consent them and welcome them onto this protocol. Then the protocol begins. Tim mentioned the tumor harvest, and he fielded some questions on that. And so patients, if benestac disease, whether it really depends on intensive discussion of myself and our multidisciplinary team, including our surgeons who are integral if the tumor, in this case, colorectal cancer, has gone to the liver, then we call upon our expert liver surgeons. If it’s gone to the lung, then we call upon our expert thoracic lung surgeons. And if it deals with the primary tumor or other areas, we call upon expert surgeons. But when the surgery is performed, we hold chemotherapy for a bit to make it safe to proceed with surgery. And then the surgery is performed, and then, as we say, we transition from the clinic back to the lab. And there’s a large, vast team that does this. And it’s not just a wet bench lab, but it’s also a core facility, molecular and cellular therapeutics I mentioned earlier, led by Dr David McKenna. And this is one of five NCI sponsored such facilities in the nation, and I think that’s. One of the aspects that makes our program, our university and this trial unique is that rather than having to send this to a different site, we do this at the university, basically on the premises, and so in collaboration with Dr Starr and Dr maherdi and Dr Weber Dr McKenna and their vast team members, there are different things going on at different points of a next month or multiple months, however long it takes to be able to derive TILs, hopefully successfully and hopefully robust. I noted a question about lymph nodes. And lymph nodes tend to be scant and couldn’t be necrotic. But the key here with infiltration is you need enough tumor tissue, and there has to be enough size there. So usually solid tumors in the we call the organ parenchyma, inside an actual organ are usually ones most likely to yield enough tills to expand they undergo genomic sequencing, and much of which I appreciate from Tim so I don’t have to expand upon this too much, but then also being able to grow them to enough numbers and also energize them with the candy that Has il two and of course, a CRISPR assist Knockout is an essential part of that process that makes this trial unique, above and beyond the studies that were done in the NCI. And then we go back into the clinic. That’s a proverbial clinic, because much of that happens at an inpatient hospital stay that we’ll go over. So the overall schema is that when someone is admitted into the study and meets eligibility for safety. They undergo apheresis, they undergo tumor resection, and we do the leukapheresis prior to the resection, and also later on, about six weeks after till infusion to compare, and also just to ensure that the tills are persisting, primarily because want to make sure that this is not just something that’s going to last a few days, a few hours, a few days or just a few weeks, but rather be a long lasting form of treatment, if it’s going to work, and hopefully will persist over time and be very efficacious long term. That is the intent, or the intent to cure, is something that’s an ambitious goal of this, but trying to go for something above and beyond the normal standard of care, such a cytotoxic chemotherapy, and even even in combination with biologic agents that are not capable of eliminating or producing a cure or completely removing tumors, rather, are able to achieve stable disease for a prescribed period of time before tumors evolve and eventually become human resistant. This is basically verbiage that describes what I showed in the schema, and the key is, when someone recovers from the tumor harvest surgery, they can receive the protocol allows for either no chemotherapy or standard of care chemotherapy, and that can be comprised of any standard of care FDA approved drugs, such as fall Fox full theory, combination of five fu based therapy with or without biologic agents or other targeted agents. And that is done in consultation with me. And if, obviously, if someone doesn’t live in Minnesota, we don’t bind them for that several month period to live here, but whether it’s winter or summer or not, but integral to all this is close collaboration between me, my entire team, and the home oncologist, working together, and that we welcome that person into the team, the local oncology team, but the imaging for disease response has to be done here, because we want to make sure it’s uniform and get as accurate as read as possible. For patients who want to know, did this treatment actually work once the autologous sis knockout, till product passes final lot release, inquire preserved the treatment with tail product maybe may go forward. So it’s not going to happen in the exact day, because some of it’s unpredictable. It will take some time to recognize the antigens to expand. And then we also have to coordinate many, many moving parts, among which is, when does the patient start chemotherapy safely after the tumor harvest, and when can they stop? But a key to that, like almost pretty much any clinical trial you can imagine or have seen, a 28 day or, in other words, four week washout is required of this trial, as it is standard in so many clinical trials, before beginning the lympha depletion chemotherapy. And again, that part of that is a lot of logistic working out with the oncologist, or if they’re under my care here, that I work that out with my team, and we’re very cognizant of that. So leading up to that, after the minimum 28 day washout comes the time which I call the centerpiece of the treatment, which is actually getting the tills. And I think, if I don’t get to say it outright later, the centerpiece, the absolute centerpiece, what we call Day Zero, literally, is the infusion of tills. And the tills is not a repeat administration. It is a single, one time administration for everything leading up to that one day and everything to follow that is surveillance to determine safety and efficacy is not going to be a repeated administration. This is intended and built to last, and that is the great hope of what we’re trying to discover on this trial. But if we call that day zero retroactive to that seven days prior is when a patient’s admitted, it may not be on a Monday, but just to make it logistically make sense as an example, if someone were to be admitted on Monday, they may receive five consecutive days of chemotherapy, which is high dose, and these drugs are things that are probably not familiar to you, because the average someone, anyone caregiver or an actual patient who receives treatment with chemotherapy for colorectal cancer may not see these unless they know someone with leukemia or lymphoma. But these are part of the. Basically the same borrowing from the same regimen, the same drugs and same dosing and schedule and frequency as what was done at the NCI. And so this is done in in supreme collaboration with our bone marrow transplant colleagues. The University of Minnesota has over half a century of experience in adult and pediatric bone marrow transplant, a very prolific and esteemed group, and they have partnered with me and Dr Jeff Miller, who I mentioned as my co investigator, is integral to that for safety when we admit these patients to our bone marrow transplant unit, but the multidisciplinary team is taking care of them, but the day to day to the doctor who rounds will be a bone marrow transplant specialist, because they have a lot of experience with these type of high dose chemotherapies. And seven days from admission, if all goes well, then we anticipate day zero would be the infusion of the tills, which is in some measure almost anticlimactic, can take an hour, it can take two hours, but a couple of hours, and then what happens at that point later in that same day, and the several days that happens after is up to six doses of the high dose, il two. It’s also called aldose Lucan, if you end up googling at home and it’s we do safety intensive safety monitoring to see if it’s safe to administer the IL two because certainly there are potential for strong side effects there. But again, the intent, just like it is in the lab, is to boost the energy of these teal cells once they’re infused into the human patient, and to make sure that they last for a long time. So by day 28 so we asked them, approximate amount of time that someone might be in the hospital as a very gross estimate, maybe two weeks, but it’s definitely not a couple of days and someone goes home, it could exceed 14 days. It may be slightly under 14 days, but I think 14 days is a good frame of reference. And when someone is discharged safely from the hospital, following this, then we have a very close follow up in our outpatient clinic, hand in hand with our bone marrow transplant and solid tumor oncology colleagues. So the patients will be seen twice a week in our outpatient clinic here at the university, and then until day 28 and day 28 measures 28 days plus or minus one day since the infusion of tills, and we will obtain scans and we will do a six week non mobilized leukophoresis to help to detect and confirm persistence of the tills. And then we will do CT scans, or other forms of scans, that might be appropriate at close intervals for the several for several months, and then spacing it out every several months after that, per our protocol. And so at the time, if disease progression occurs, then I consult with the referring physician, or if they’re under my care for anything appropriate. But we would like to continue long term follow up study again looking for long term follow up of safety as well as efficacy. There’s a long list of inclusion criteria that are fully available on the clinical trials.gov so I won’t touch on every single one, but probably the things that people want to know in general, definitely for a metastatic gastrointestinal epithelial cancers, patients who’ve had progressive disease after at least one line of therapy. And so that first line of therapy, for example, if Colorectal cancer is diagnosed and a stage four diagnosis, just as one standard of care, example might be FOLFOX with Avastin, if someone has progression disease on that would potentially be eligible. And I would love to hear from patients at that stage. And then maybe someone has received two lines of treatment. And that’s also an opportunity to inquire about eligibility, if it’s not for pediatric patients, and it’s not for patients who are elderly and maybe have a poor performance status, but 18 to 70, as per the trial, to respect those parameters. And you know, standard for any clinical trial hematologic and adequate liver and kidney function of the organ function, we want to look at more than four weeks elapsing from prior systemic therapy for that 20 day washout period. And respect to the fact that we’ve we’ve encountered many inquiries, not just from Minnesota, around the world, around the country. We understand and plan for the fact that not everyone on this trial will be from Minnesota, per se, but for the first 28 days, at least until the end of treatment, visit, patients must agree to be remain within a one hour drive of the University Minnesota. I would say, we want to work with patients and partner with patients, but I would say that’s not something for safety reasons that we could change. It has to be something that someone has to be willing and so one of the early questions that I trained my team in taking to ask is if someone feels that realistically, they would not come to Minnesota for more than a few days. And I say this is definitely not something that would be appropriate for them, or maybe not be of interest to them. So due to a 10 to 12 week or more delay between study enrollment, start of study treatment that could be variable based on the tills, we have to reassess performance status, just to make sure, again, for safety purposes, that someone is healthy enough to proceed, that hematologic parameters and blood count and other organ function, as demonstrated by labs and scans, is safe to pull forward. There’s a long list of eligibility, and also just ensuring that people don’t have other types of viral infections that could make til infusion or the lympha depletion chemotherapy in particular, more dangerous. Just ensuring that women with child bearing age are not pregnant, and just testing for that any changes in medical status or. Social situation that would make study participation not in the best interest of the patient and the opinion of the role investigator. So some of that might be if they have interim hospitalizations, if they’re extremely ill, may not be good time. But as Tim mentioned, if someone were to cover the tills are frozen and could be thought if appropriate, at a later time and in social situation. I think honestly, just for as many bone marrow transplant candidates, it’s an intensive process that requires a lot of social and family or caregiver of any kind support. I think that holds here, and it’s not something that any individual can physically or in many cases, do alone, but would need help. And so that’s something that would be really important to help arrange continuing to agree to remain within the Twin Cities area for 28 days, as I mentioned, and for informed written consent for any clinic trial. This is a brief schema. This is showing in schema format, the number of days what chemo therapies will be given. And this is the example, Monday through Friday. I gave cyclophosphamide given on the second and third day, but fludarabine is an intensive drug given five days in a row, started the day of admission, a little bit break of two days, and then anticipated till what we call Day Zero, followed by il two for up to several days, but a maximum of six doses. So I think vigilance is essential for safety. And so to answer some of the questions that people will naturally have upon hearing of this trial, there are a couple of novel areas, and with great novelty and innovation come responsibility to to understand what the potential side effects might be, some of which we know, and many of which from the NCI experience, can be attributed to either effects of the aisle, two such as high fevers or rigors or chills and the level of depletion chemotherapy, which essentially brings its goal is to bring the bone marrow and wipe out the existing immune system, to replace it with the immune system supplemented by the tills. I anticipate you may have, as many people might have, also in the news read about the so called cytokine storm, also called cytokine release syndrome, that has been associated mostly with car almost all with car T cells as an example. FDA approved for pediatric and some forms of adult liquid malignancies, but not found to be an experience to date, particularly since all that seems to be at the NCI. — but not seen and experienced, a date at the NCI with tills or another centers, nearly, pretty much the entirety of expected, . The entirety of side effects are anticipated are from the high dose lymphodepletion chemotherapy in il two that’s something that does differentiate this form of therapy from adoptive cell therapy off so called off the shelf car T cells, of course, we might discover because we’re at the cutting edge of innovation. Sis, immune checkpoint abrogation with CRISPR, cast nine produces one or two factors that we’re going to discover whether or not that rule might be different, or if that will hold for T cells that have been altered in this way, but we will do very careful monitoring of sign, of sign, vital signs, and these are some of the potential side effects of cell infusion that we want to look out for, and others. So I emphasized earlier that patients will receive a single course of treatment, including live with depletion chemotherapy, followed by the tills and then the aisle two, up to six doses. But we have to predicate on safety, and so I say up to eight doses of aisle two, and that might be one, that might be six, it might be three, might be zero, if it is not deemed safe. And that’s me on the premises constantly, every eight hours, getting calls from our inpatient team to assess that, as well as in person assessment. And then a treatment visit occurs on day plus 28 there’s a long list of assessments that, of course, to understand safety and to be vigilant about that. I said vigilance is required for safety. There’s a lot people will get to know our staff in the clinic, in our hospital, and through phone calls, a lot, including virtual visits in this day and age. So we have intensive mechanisms of reporting for governing bodies that ever incident monitoring and documentation through our Clinical Trials Office, we have overall experience with cellular therapy trials. And so this is just the new wave of evolution for that. And so having gained the experience, having opened the trial in the past year, in 2020 we’ve already gained a valuable experience, and also in close collaboration with a lot of colleagues who have experience at other centers. So I think from a logistical standpoint, some people may have questioned about, how would I find out more? We funnel the input system for patients not already at the university, but certainly, if patients are already treated at the university, can find out more or in the region or in the world or anywhere else in the country. You can look up clinical trials.gov NCT number, and we are funneling the inquiries to our expert clinical trial nurse navigator, who’s on the other end of these phone numbers, the phone number and the email. And so I appreciate sometimes people want to find my name or find other. People’s name and look up because it’s easy to do that Internet, and I appreciate that. What I don’t want is for referrals to get lost in the process, and also, in some measure, some of it is not HIPAA compliant by standard email. We want to respect privacy and ensure that. So the best mechanism I could recommend, and this is the same information copied from the clinical trials.gov website, is this phone number goes directly to our clinical trial nurse navigator, specifically for this trial and others, but primarily for this trial. And this is the email address. So I think of having worked with the ACR scientists survivor program for better part of the last decade, I’ve really come to have extreme admiration for patient advocates for a few years as yourself, because I know it’s not easy. It takes time. And I think, you know, having had loved ones with cancer, I feel like it’s almost a full time, or at least a part time job, just to navigate complex health systems, to navigate treatment for yourself, and deal with insurance companies and so people who take time out of the normal life, so to speak, to do this. I don’t know why I do it, and I have, I think, of immense courage. And the fact that even want to know this level of detail is never ceased to amaze me, but it shows the vast investment. And there’s nobody who has better investment than patients and their caregivers to find something better than what already exists, to find the next generation of cures. And so when patients volunteer for clinical trials, I’m always just left just very with very much with profound admiration. I’m gonna pause there. I don’t want to get too emotional on that, but I think bring cutting edge of care is the dream of scientists like Dr starr, Dr minority, Dr Weber, physicians like me who can help bridge that this, this is, this is something we’ve all dreamed of. We didn’t know about CRISPR cast nine until exposed and invented to the world in the last decade. But things like this are why we do cancer research. People like you are the reason we do all this. And so whether it’s an endeavor that takes up our days and nights and we lose sleep over we know that the end goal is to really do this, and it takes a village, and we want to bring it with great safety to everyone. I will bring over here. If you can see, this is the clinical trial.gov page. This is the information at the bottom, saying, recruiting. If any one of you has interest in the actual references, some which Tim alluded to, you can click on here and you can be taken that set of information, the long list of eligibility I basically took verbatim here, so it’s all included, and also any more information. The basic description This summarizes nicely the essence of what I’ve described. And it’s the same title that I described on the title slide, so I know we have just a few remaining minutes together, and I’d be very happy or to tag team with Dr Starr or any appropriate questions that I can answer.

Manju George 47:45
Thank you so much. Excellent. That was excellent. That was a very good introduction. You know, great introduction. And thank you, Doctor starr. I mean you your your explanations at the stage. So I think that like for us in our group, because we have the til trial, and, you know, we have people who have been posting about what they’re going through, like, the whole chemo regimen, and what they have experienced, and when they got the kills, what happened, and when they got the ILQ, what happened. So I think that lots of people maybe on this call may be familiar, because, you know, this is an advantage in having these groups right, that they explained like so this, you know, so I hope that the listeners were able to kind of place what you spoke in the context of what they’ve heard in the group. So for me, I think there is some concern about the chill trials in our group because of, you know, the cytokine storm and things like that. And, you know, one of the things I think people feel very comfortable is that they would be at NIH and it has the, you know, that they would get a lot of care and all of that. And I think that the way you have explained with the your university’s experience in bone marrow transplantation and all that you it looks like you have a similar team and the same setup and all the monitoring and everything, right? I think in my mind, that’s what I’ve heard the most common concern that, you know, how safe is it? And I mean, your study is exactly looking for that, right?

Dr Lou 49:10
I’ll tell you, I never set for the state of Minnesota until I interviewed for position here, and lo and behold, here I am, and that’s a decade later. So, you know, I think maybe allow me just provide some context so I have the background here. So there were approximately, I think now, over 60, if not more, National Cancer instituted designated comprehensive cancer centers. That’s basically a designation given for cancer centers of excellence and academic settings that have the full spectrum of Premier clinical care as well as cancer research and things in between. And there’s, there’s a large application process every five years to get this gigantic grant to have this designation. And the Cancer Center, which is called the Masonic Cancer Center at the University of Minnesota, has been an NCI designated Comprehensive Cancer Center for many decades. And there’s a specific reason. It’s not that. By accident, and it’s not only because of the basic science research here at the university that we have this trial. We were brought the opportunity to have this trial in partnership because of the renowned and because we had everything under one roof. So it was gi oncology expertise. And that’s not just me, obviously, it’s a whole team of people, and that includes expert surgeons, oncologists, radiologists, radiotherapists every everything, and then the basic science also, and then also the Molecular Cellular therapeutics group, which is one of only five NCI sponsored groups in the country. And you know what? The BMT group is one facet of that, but it’s crucial. And I think something that’s been that we were told, and one of the reasons we were asked to take on this trial is that the NCI is, you know, my best friend’s mom died of pancreas cancer in the early 90s, and that was a time where that was a last ditch effort. But in the NCI has certain capabilities, and certain capabilities it does not have. Is it premier center, and a lot of great research comes out of there, but they’re not a tertiary care referral center. They are there. And many times, for patients who have exhausted all available options and nothing else available to them, or cannot pursue anything else, they might go the NCI. Whereas here at the University of Minnesota, from the beginning, when someone is first diagnosed, when a colonoscopy is first performed, here or at our affiliate sites in the region or throughout the state, they are diagnosed colorectal cancer, they have access to our Clinical Trials Network. They have access to us. They can meet with me when first diagnosed, and maybe for the right patient, I might meet the right patient and think this patient 612, months down the road might be a good candidate for this trial, because we’re running this trial here and all the way to enrollment. We have not just the basic ability capacity. We have the outstanding capacity to take excellent care of these patients. So I think I just want to reassure people, maybe are not familiar with the state of Minnesota, not familiar with the University of Minnesota. I understand that there’s a at least one other center that may be more famous, but we have this trial. And I think there’s a reason why we have this trial, and not to say other centers won’t in the future or not, can’t or don’t have great trials themselves, but this is an innovative trial, and I think it’s because of our expertise as a multi disciplinary team that we have the opportunity to do this. And it’s a great undertaking. It’s years in the making, not just the basic science, it’s years to get the point we could even open the clinical trial, years of clinical work to get to this point, we could do this safely.

Manju George 52:22
Okay, I have one question, Would you be open to having tele, tele health visits with patients? Because I can imagine that after they watch the videos, they probably want to have, you know, initial consult with you before they, you know, decide to come. Is that something that you will be interested in doing?

Dr. Starr 52:38
I’m certainly interested. I have to work within the boundaries of what I’m allowed to do within our health system, as in many, nearly all states. Well, we, with the onset of COVID, a year ago, we plunged into the world of virtual medicine. I’m doing that almost exclusively. I still encourage you as a first point again, funneling to the clinical trial nurse navigator, the phone call and also the or email, because what the trial nurse does is she helps gather all the information. I can’t make an assessment video or not, wherever someone in the world is without proper information, and our team meets multiple times per month, led by me, but our whole team to go through scans, to go through information. I spend a great deal of my time, day and night, doing that, to try and not waste people’s time at the same time. Practically, we are not allowed to do virtual visits for people in other states, aside from Wisconsin. So unfortunately, although you think you know FaceTime and zoom and HIPAA compliant versions of virtual visits break down boundaries. Legally, I’m not allowed to this is a state government level. It’s not our institution even to to do that unless someone is physically in state of Minnesota. I acquired a a temporary license for the state of Wisconsin for the remainder of the year in order to take care of patients, my patients, and other future patients who live in Wisconsin. So we don’t have to physically come over. But if someone lives in Hawaii, for example, unfortunately, I’m not able to, but I’m open to discussion, because I don’t want to waste people’s time and have them come over here. But we do a lot of homework behind the scenes, but that’s starting by spending a lot of time obtaining the records. So I would encourage you, if you have interest, please don’t reach out to me directly, not because I wouldn’t love to hear from everybody, but we have a system in place to ensure we don’t miss any information, and my team helps me do that. Okay,

Manju George 54:22
that makes sense. And then you said that the dose of the til was one time. Are you guys also planning -??- own cells, and is there any any plan later to be using them, or something like that?

Manju George 54:39
I was wondering if you will be also freezing down cells in addition to infusing

Dr. Lou 54:46
so we are the administration clinically for the patient on the trials. It was intended to be one time for the current time, maybe future iterations of the trial might do different. But we are also doing extensive research in what we call correlate biomarkers, and that effort is led by doctors. Are. Our whole team is doing that with blood and tumor samples to determine, once we have the clinical outcomes and see who whose tumors respond best, try and identify specific factors of why that is, and maybe to refine the trial further. But for this iteration of trial, the administration of the tills is is intended to be one time. It’s intended to be hard hit, however you want to call it unintended, for this to be built to last and to have a durable, effective response while also being safe.

Manju George 55:27
So my other question is, thank you very much for that answer. My other question is, on the clinical trial website, you have a list of mutations listed right? So Should people with only those mutations reach out to this clinical trial coordinator, or is it?

Dr. Lou 55:45
There are some team mutations and the chaos mutations and all of those listed. Yeah this is independent. So that’s a great question, because so much of in the era of molecular profiling and molecular oncology basket trials, or umbrella trials, whatever we wish to call them, predicated on a tumor harboring a certain mutation at a certain time in tumors evolution. This is independent. So it has nothing to do with Pdl, one if you’re if you’re interested or read about that is nothing to do with micro satellite instability, presence or absence of K RAS mutation, BRAF mutation N RAS is independent. So it’s really more predicated, as Dr starr said, on the new antigens, and being able to stimulate the new antigens to be productive and be recognized, but is independent of any of the status that we normally would identify on standard of care genomic profiling,

Manju George 56:32
okay, I think I see one question here. It is like, can you comment on vitals without the CIS blockade are not very effective in solid tumors.

Dr. Lou 56:44
Well, I think the numbers, we don’t have 1000s of experience, but some of the studies from the NCI, I mean our limited number, and I think logistically, these are very, very difficult to try out, to carry out, and they’ve been step wise over time. So it’s not that they don’t respond without sis knockout. But this effort to knock out sis is an effort to improve further upon efficacy. I mean, the golden prize. We would love it. We would love it if 100% of time we did a treatment that it worked, we know that that’s not the case for any form of targeted therapy, regular chemotherapy or immunotherapy or cellular therapy yet. But if this strategy works, would sis be the next level? Would it improve? It double or triple fold? That would be great. So I don’t think it’s a till that don’t work, but really it’s a first step, and what probably probably be many steps in the years and decades to come.

Manju George 57:34
Okay? Thank you. So I think Dr Star had answered about the size of the tumor. He said about 10 grams. So how much like do you? Do you have? Like, I know it’s not like a fair question to ask, but what size match would be approximately? Is it like five, six centimeter cube lesion, or do you have any? Ideally,

Dr. Lou 57:55
I would say, as much as safely could be possible, but very grossly. I won’t stick to it exactly, but like, if you imagine a centimeter or something in centimeter or more in size, when I connect with our surgeons, we look at it and it really it’s kind of a game time decision, because a lot of time tumors that are treated or grow rapidly can become necrotic, meaning they die, which is good. It could be because of tumor so large that the interior part dies, or because the chemotherapy has been so effective, but if the cell portion is dead, it doesn’t have the tills that we can extract. So in that case, we may need to extract more tumor, and we also want to do some for research purposes as well. So minimum, I guesstimate, like a centimeter, and we try to get as much as safely can be removed. But also, just as we round up the discussion and appreciate your time. Is that the surgery for tumor harvest is not done with intent to cure or remove the entire tumor, but rather with intent of getting enough to make the till extraction feasible. I appreciate your time. I do have to move on, but I’m very grateful for the opportunity to come meet with everybody. Thank you, Manju, thank you. Thank you. Dr star for helping me out. Dr George as well,

Manju George 59:03
yeah, thank you. Dr Lou, thank you. Dr Starr, this is great. Thank you very much.