Founder Stories: Heinrich Haas, RNA for Individualized Cancer Immunotherapy

Speaker, which is Heinrich. So is he here?

Oh, very good.

We'll hand it with your capable hands.

So, ah, okay. Do we have a pointer or— this one here. Okay. And how do I proceed with—? Okay, so thank you very much for the introduction. Thank you all for staying so late, and thanks also to the organizers for inviting me to give this presentation on our developments in the field of tumor immunotherapy. The title of my talk is RNA Nanomedicines for Individualized Tumor Immunotherapy: Translation from Basic Research into Clinical Phase I Trials. And therefore I think I meet some of the keywords which are required for this conference. And before I go into detail, I would like to present to you our organization, which is a little bit particular. And before I present the organization, I would like to present to you the key persons who more or less were fundamental for having this going on with their research at the medical school at the Mainz University. Christoph Huber, Özlem Terzi, and Uğur Şahin, who did groundbreaking research and then initiated certain spin-outs out of the university. First, I founded Ganymed in 2001, where we developed monoclonal antibodies, so-called ideal antibodies. BioNTech, the company where I am in, was founded in 2008. There is a kind of nonprofit organization, Tavon, which is functioning as an incubator, as an intermediate step between academic research and pharmaceutical development. There's KIMT, which is a conference on immunotherapy, which takes place every year in Mainz. And we won a grant for an excellence cluster, also focusing on individualized immunointervention, which went to the whole Rhine-Main area where we are also in. I'm at BioNTech, and to give you an introduction into BioNTech, which is also a bit complex, BioNTech is a holding consisting of several daughter companies which all follow individual concepts focusing on therapy or diagnostics in the immunology field in the widest thinkable sense. I would not like to go into details. What I will present to you is results from BioNTech RNA pharmaceuticals, but I would like to mention that we do have a manufacturing site in a city close to Mainz where we manufacture our RNA and the liposomes we use for our investigational medicinal products. So, tumor-immune immunotherapy. As you all know, and it has been mentioned several times during this conference, tumor cells can be discriminated from healthy cells by certain motifs, by tumor-associated antigens, and these antigens are recognized recognized by the immune system. So the immune system efficiently kills tumor cells, so that's why not much more people suffer from cancer. But in certain instances, it is not able to attack the tumor cells anymore, and therefore our aim is to activate and selectively direct the immune response, the natural immune response of the body against tumor cells via the tumor-associated antigens. How do we do that? We do it using messenger RNA. So we do not deliver conventional or classical antigens like in a vaccination, as we all know. We deliver messenger RNA, which codes for a tumor antigen. The messenger RNA is delivered to antigen-presenting cells, and this initiates a whole cascade cascade of immunological processes where in the end proliferation of tumor-associated antigen-specific T cells is stimulated. And these T cells are supposed to attack and kill the tumor cells. In addition, we have several other immunological responses which act in an adjuvant manner towards this immunotherapy. So how do we get our RNA? Our RNA is manufactured by an enzymatic process based starting from a DNA template. And the whole process of RNA manufacturing has been optimized towards improved stability and expression efficacy of the RNA. So we have virtually optimized all kinds of molecular functions in the RNA, from 3', 5', UTR, the cap structure, the poly-A tail, and from that we ended up in a highly efficient and long-lasting expression of the antigen or the protein from the RNA. One key reason why we do this is that with the RNA we can go into real, true personalized medicine in the sense that we start off with a patient. The patient comes to the clinic with a tumor, a biosection from the tumor is taken, the tumor is sequenced in comparison to healthy tissue, and then using certain computational algorithms, we identify certain mutations that code for epitopes, and we identify these mutations that are most favorable in the sense of an immunological intervention. Having identified these epitopes, we translate them into a DNA template, and from that the RNA is specifically manufactured for this individual patient under GMP. And then we have the drug product for treating the patient ready in a timescale of weeks. Actually, the longest time in this process is the sterility test for the final drug product as listed takes some weeks. Now we have an individualized drug product and we still need to bring it into the patient. And for bringing it to the patient, delivery is the key word. And we have two main routes of delivering RNA to antigen-presenting cells in the patient. One is a local administration into the lymph nodes. Where the RNA is injected in a buffer system and it's uptaken in this confined space by macropinocytosis by the APCs. This is a highly efficient and effective way to deliver RNA, and we are devoid of all the problems of delivery or degradation by RNases, serum interactions, and so on. However, to be more versatile, we have also developed a platform a lipid nanoparticle which can be administered systemically, intravenously, and which then migrates selectively to the organ of interest of— and here we are in the development cycles, we were interested in the spleen because spleen is rich in dendritic cells, antigen-presenting cells. So for the further talk, I will focus mostly on these formulations, nanoparticle formulations for systemic delivery after systemic administration. Here I show you what our formulations do. These are two different formulations of RNA. RNA is coding for luciferase and we do bioimaging. And you see that in formulation one, we get preferential expression in the lung and a little bit in the spleen. While for formulation two, we get almost exclusive expression in the spleen. And as the spleen is our reporter organ for antigen-presenting presenting cells. This is kind of a target formulation for us for further development. So I think we don't have so much time to go into details, but I just would like to give you some snapshots of how we came to that. Basically, our formulation development was driven by physical and physical chemical insights, just like Amalda Ferrari presented yesterday. So we look on the nanoparticles as colloidal systems which are characterized by parameters like size, phase state, state of molecular organization, rigidity, and then correlate physical chemical parameters with the performance in vitro and in vivo. And here is your typical curve which we measure, so here we incubate RNA and liposomes, cationic liposomes in that case, in different ratios, and we measure particle size, and you see there are ranges where the particle size is confined, where it diverges,— this is a charge equilibrium, so you screen the electrostatic repulsion, and then it goes down again, and in the same time, the zeta potential switches from negative to positive. So we can make these particles and we can bring them into biological testing, and we look also on many other parameters here, just to give you an idea. I don't discuss this data. We can extract lots of information on them here. You see X-ray scattering data, you see nice peaks, which correspond nicely to the information we get from electron microscopy in terms of lamellarity and internal organization. And based on this data, we test certain systems in vivo. And here you can see how we now can modulate, we can steer, we can direct the delivery from one type of organ to the other. And as a result, we select the one which is optimum optimum in terms of efficacy and selectivity and take that as a basis for further clinical development. So we had our clinical development candidate. To come up with a, with a clinical formulation, of course you have to do upscaling, you have to change your manufacturing protocols for both the precursor components and the final IMP, which we have done. We found GMP-compliant methods for manufacturing of all of our components.. And in the end, we came up with a kit approach for our investigation medicine and product where we deliver RNA and the diluent and liposomes in separate vials for preparation of the injectable product directly prior to administration to the patient. The reason is that this protocol is really versatile and allows us to use it for for many different covariances in different clinical studies. The constitution of preparation, this actually depends on the opinion of the local authorities. It's very easy and straightforward. You dilute the RNA with a diluent, and then you add the liposomes, and you end up in a slightly opaque dispersion, which is then ready for injection to the patient. Here I have listed some of the antigens which we use for our first clinical studies. Some of them are well known from earlier cancer vaccine trials. Some of them have been tested already in earlier studies from ourselves where we had intranodal injection. And there are also novel targets. And the important thing is that we combine 4 different antigens which are extranuclear and what's to the patients. Tox data were nice and unproblematic, excellent safety profile. We found pharmacodynamic effects which were typical for a vaccination reaction, like certain cytokine levels, but which went back to the ordinary level soon after administration. This is some information on the first clinical study we initiated with these 4 tumor antigens I mentioned to you. So we started just this year in May and treated the first patients already for 5 cycles. We continuously increase the dose and found already at relatively low doses indication for for immunological responses, which is good in a certain sense. And so we hope that this will continue to see— give also information on the expression on the direct antigens, which we have not measured yet so far for this study. Here, this is a summary of other studies we have initiated or which are currently in planning. There are studies with intranodal administration administration as well as with intravenous administration. There are studies with generic compositions of definite antigens. There are studies where we use a so-called warehouse where patients are stratified and from the warehouse the most relevant antigens are selected and we have strictly individualized medications which are administered to the patients. This was the end. I would like to thank all the collaborators. This is a collaboration of many different people in the company. This is preclinical department: Lena, Kerstin, Mustafa, Yves, Sebastian, many more. Andreas Kuhn and the RNA biochemistry group. The project managers, here just two of them. And this is some people from the formulation development group. Who worked on the formulation development, Gerald Placisinski from Max Planck Institute of Colloids and Interfaces in Golm, and Sergio Finale, where we made some of our extra measurements. And of course, nothing of this would have been possible without Ugo Sein, who is the great driving force and who is really providing a stimulating environment to bring this ahead. Thank you for your attention.

We're going to reset the clock now. We're going to reset the clock now. And oh, it's going. It's going. There you go. Time for 5 minutes of questions. So please.

Hi. Just a question about the choice of the intravenous route for administration. In some many cases, RNA lipoplexes have been found to be toxic by the systemic administration because of these cytokine releases and other other systemic toxic effects. What is going to be the advantage for a vaccine by being given by the intravenous route as opposed to subcutaneous route? Yes, that's a good question. Of course, we have tested also subcutaneous, intramuscular, and several other routes, but we found that with the intravenous route we get this very high selectivity which depends on the particle properties. And one big advantage is that we can address much more antigen-presenting cells compared to an intramuscular, which is kind of an almost local administration or sub-q administration. So we have some technological advantages. We have more parameters which we can vary for future development. We can administer more or higher doses. And so we have more freedom to operate with this kind of administration. Okay.

Question there, please. Yeah. Thank you very much for the talk. My question goes also in a similar direction. So it's not clear for me what is the selection, what is the individual component, because you said that you are selecting some antigens or epitopes, but it seems that this is a universal vaccine. So do you have some particular peptides of all 4 antigens, or are 4 antigens out of many? And then, do you use any adjuvant because the IV is not the most immunogenic route that you could imagine?

Yes. So we, as on the first question, we do all of it. In some of our studies, we have fixed composition of antigens which are known from the literature or which have been developed by us. In other studies, we have the so-called warehouse approach. So this is for the TNBC breast cancer study. Patients are screened and we have a warehouse of 20 to 30 antigens, and so they are administered with the antigens for which they are most positive. The selection is also done by certain algorithms. For the personalized approach, this, as I said, We sequence the patient and apply algorithms in order to identify the most immunogenic epitopes. So it's really strictly personalized for the patients.

What was the second question? The adjuvant. But the most immunogenic based on what? Because if this will be extracted, this should have already an immune response.

Well, this is what the guys implement into their algorithms. And the immunogenicity, the RNA is a strong adjuvant, so we don't need adjuvants. Okay. I'm wondering about this biodistribution of the different particles. In one case, almost everything ended up in spleen. Do you understand why it's almost everything ends in spleen and nothing in liver? Yeah, that's a good point. I think the reason why it not goes to the liver is the selection of our lipids, because they are not good transfection reagents for the liver. The reason why it goes to the spleen and nowhere else is, let's say, the uptake of mechanism. Basically, as I showed you, this charge scale and what we administer is nanoparticles with an excess negative charge. So these are actually dysfunctional transfection reagents. Nobody would administer negatively charged transfection reagents because it doesn't transfect ordinary cells. However, dendritic cells and related ones, they are professionally uptaking foreign compounds and maybe particularly slightly negatively charged ones because bacteria are a bit more negatively charged than our cells. And so it is the particular uptake mechanism of the cells we are targeting. We don't need to breach the cellular membrane. It's an open door, and they catch them. I also wonder if the size were the same for the particles homing in spleen and those where you had the considerable amount in heart. In lungs. In lungs. Yeah, we tried our best to make them equally in size. Maybe there were some tens of nanometers in difference, but they were equally in size. But what I can say is that size matters. And we find that, as it's known in the community, that for this purpose, larger particles perform better than smaller ones.

I'm going to use the chairman's prerogative to ask one last question. If you could be very quick on the answer perhaps. Your dose size is quite small at a milligram, and I was wondering whether in man you're going to see a problem with natural antibodies taking out quite a large percentage of that milligram material. And my rider on it also was the cost of treatment in man. Do you have any thoughts on that? [Speaker:DR.

FRANZ SPOHN] Yeah, that's— for the antibodies, we don't know yet actually because we haven't investigated that. Yeah, that's very remarkable that we get a signal already at relatively low doses, lower than in the corresponding doses in mice. And the cost, of course, that's a key issue, particularly for the individualized nanomedicine, because you have to manufacture individualized nanomedicines, and we think we will not end up as dendrions. So our aim is to bring down the cost of manufacturing by several orders of magnitude just due to scaling and parallelization effects. Just like a USB stick or a cell phone costs nothing anymore, almost nothing, we think that we will be able to bring down the cost for manufacturing this individualized, uh, therapeutics, for example, by automatization, taking man out and having machines doing the job. Thank you very much.

Thank you, everybody.