Hi everyone and welcome. My name is Francesca Bijon and I will serve as your moderator today. Thank you for joining us for today's webinar, how to bring your lipid nanoparticle formulation to clinical commercial manufacturing via ctdmo services. As your moderator, it is my role to ensure that we make the most of your time with us. I'm here today with Mahesh Karva, associate director, process development. Mahesh leads the process development group at our lipid nanoparticle center of excellence in Indianapolis. He holds a PhD in nanomaterials chemistry and has more than 15 years experience in the Pharmaceutical industry, having worked and contributed to the success of several client projects in the NMP and liposomal fields for RNA delivery. Here with us today is also Teresa Dent, strategic marketing manager. Theresa holds a PhD in Biochemistry and in her current role she is responsible for managing our lipids portfolio and custom manufacturing businesses, supporting top industry players and young startups in the field of LMP drug delivery, Evan worked as product manager for synthetic lipids for several years. She gained valuable experience in product and process development of lipids used in lipid based RNA delivery. Before I tie things over to our presenters, I'd like to cover a few housekeeping items. At the bottom of your screen, there are multiple application widgets you can use. All the widgets are resizable and movable, so feel free to move them around to get the most out of your desktop space. You can also find our reaction button, indicated by the tonsap emoji, which allows you to give immediate feedback on the presentations or topics that stand out. You can expand your slide area or maximize it to full screen by clicking on the arrows in the top right corner. If you have any questions during the webinar, you can submit them through the via the Q&A widget. We will try to answer during the webinar, but if a more detailed answer is needed or if we run out of time, we will answer later via e-mail. Please know that we do capture all your questions. We will also have the opportunity to participate in a couple of quick poll questions throughout the session. I encourage you to take part in this. If you're watching this webinar on demand, you can still submit poll responses as well as questions. The webinar is being streamed through your computer, so there is no dial in number for the best audio quality. Please ensure your computer speakers or headsets are turned on and the volume is up so you can hear the presenters. An on demand version of this webinar will be made available afterwards and you can access using the same link that you have already received. So that's it from my side. And now it's my pleasure to turn things over to our speakers. Teresa, over to you. Thanks Francesca and welcome also from my side to this webina on how to bring your lipid nanoparticle formulation to clinical and or commercial manufacturing. We are contract testing, development and manufacturing services and before we jump right into the focus topics of today, I would like to use a moment to introduce you to our integrated M&RCDM or offering. With this, we as company can support you throughout your entire drug product development journey. Our capabilities include air and a development and manufacturing totally suitable for your customized solution. We do also offer high quality GMP lipids and there you can benefit from on the one side a broad portfolio from which we can supply. Readily available lipids that are commonly commonly used in lipid nanoparticle formulations. On the other hand, we can also support you with customized lipid chemistry depending on your needs for your formulation. Lastly, we do also offer lipid nanoparticle formulation and manufacturing services all the way from preclinical to commercial manufacturing. Lastly to highlight is that we also offer fill and finish services for your drug products. For today, we have actually actually chosen two major categories that we'd like to discuss with you. In the very first section, I will provide some insights on the critical quality consideration for libit excipients. In the second session, Mahesh will guide you through lipid nanoparticle commercialization and after some key takeaways, we will move on to the question and answer session where we will address the questions that you provided prior and during through the WEBINA. As mentioned already by Francesca, throughout the WEBINA we will share some poll questions with you and we would like to encourage your participation in order to enrich this webina also with your experience. So let's jump right into it. As some of you might be already aware of, lipid nanoparticles require different kind of lipids and those lipids can be categorized into four major categories. Going clockwise on this slide, you have the first category, which is the cationic ionizable lipids. They are responsible for the actual encapsulation of the API. So as shown on this slide, the RNA. And they are also linked to the release of the payload in our cell. The second category are then packed lipids. Those are overall responsible for the stability of the particle and the longer circulation time. Category three and four are then utilized to provide the structural support of the particle and as a consequence, its rigidity our offering both comprises. Portfolio lipids. So there we have a broad offering of different lipids that are commonly used for those different functions. Moreover, we then also have the customized offerings where we develop with new lipid chemistry and process design since we now see here that all those lipids have such a decisive effect in this delivery technology due to the various function. It is a logical consequence to highlight that the quality of those lipids is decisive when it comes to your drug development process. And if we look at what actually defines the quality of those lipids, there are different parameters that are important to consider. So the very first one you can see here on the left side is the purity of lipids. So it has been shown that. Purity of lipids has a decisive impact on the stability of the overall drug product. It has also been identified that the presence of impurities that are undesired in the lipids can, for instance, impact your drug release profile. The second pillow that is shown here is the consistency of lipid quality when it comes to lipid supply. Because of course, you can only achieve reproducible results in your drug product as well as in your validated processes if you have a consistent supply in the quality of your raw materials. As a side effect here to mention is also that if you have a consistency in the quality of your lipid supply, you can avoid the necessity of bridging toxicity studies. The third pillow that we'd like to highlight are the overall physical chemical properties of the the lipids used in your formulations. Because this is also linked to their handling properties during the process and to the material characteristics, there are parameter important to highlight, such as crystallinity, the solubility behavior of lipids, their stability and their flowability. When considering all those aspects in your process development, you will also recognize certain benefits, and the very first one that we'd like to highlight here is of course, that if you use high quality raw materials, the probability of actually achieving your desired result in the drug development process can be increased. Moreover, in these days we observe increasing regulatory requirements for raw materials as well as a novel modality drug products. And therefore we believe if you focus on the right supplier that can support your wit, both the quality and the supportive documentation, you do have the chance to better manage those regulatory challenges. Lastly since libits. Are so decisive during the drug development, manufacturing and the process design. We believe that you can facilitate this journey by using high quality lipids and thus also have a cost saving effect overall. So now we have to look at it from a more broad picture perspective and with this, we can jump now a little bit deeper into that quality topic by looking at 2 specific case studies. For those case studies, we have chosen on the one hand our cationic lipid Dota that is used for encapsulation and deliveries of nucleic acid as well as on the other side DOP a help a little bit that has the role of cell uptake and endosomal release. On the slide before we have pointed out purity as one of the decisive quality parameters. So when we. Develop new lipid chemistry purity is one of the aspects that we always keep in mind and as you can see on this slide, a comparison of purity data conducted by HPC showed that our commercial available do tab with 99.9% achieved higher purity values compared to other products available on the market. As a consequence to that, we do also then observe lower levels in undesired impurities and as they can have a decisive impact on your process performance and are strictly regulated by the regulatory limits. This is of course also something that you can benefit from, in particular when we talk about the next step, so the application. In in the formulation experiments, the handling properties of lipids is really important and therefore we also focus that when we do our product designs. As you can see on this slide, our do tap in addition to high purity is also a nicely crystalline powder and therefore enables quite good handling properties when it comes to the application as a next step. A similar comparison has been then conducted for DOP, our helper lipid and also here you can see that our DOP achieved significantly higher values when it comes to purity. One further explanation that I'd like to share with you here is that we are relying all our lipid processes on fully synthetic approaches. And this enables us to use or utilize different and various purification steps throughout the process to maintain those high levels in purity. And with this, I invite you to participate at this very first poll question. We would like to understand here what according to your opinion are the most critical quality parameter when it comes to lipid formulation. So you can choose between a the libit handling properties, B libit purity, C libit stability and ease of storage, and D libit packaging configuration. I will now give you some time to vote and then we can look together on the result. So I will read it once more for you. Select the top three parameters crucial for your formulation answer. A lipid handling properties B lipid purity, C lipid stability and ease of storage D lipid packaging configuration. OK, that's very interesting. So as also highlighted by our results, purity is one of the aspects that is pretty highlighted here. OK, so let's move on. To conclude my part of today, I I'd like to highlight again some of the key takeaways that we saw today. So it is important to note that lipids do have a decisive impact not just on the final drug product, but also the process of the lipid nanoparticles. Particularly, we would like to highlight the importance of purity as well as the physicochemical properties of the lipids. So overall, we believe that quality should be really decisive parameter when you are selecting your lipid supplier. And since we always keep in mind all the different steps required during this development journey, we are also considering the formulation, so the lipids application when designing new products and therefore we believe that. We can be the perfect partner to contribute and support your drug product development journey. As a last question from my side, we again like to invite you to vote here. It is related to understanding for which part of this drug product development value chain you are seeking support for. So please vote between custom lipid development and manufacturing. C Custom, M RNA development and manufacturing, C custom lipid nanoparticle formulation development, D lipid nanoparticle manufacturing and E fill and finish capabilities I will give you again some time. So what type of CDMO services are you looking for? A custom lipid development and manufacturing B custom Aaron. A development and manufacturing C custom lipid nanoparticle formulation development? D lipid nanoparticle manufacturing E fill and finish services. OK. Also pretty distributed. And with this, I will hand over to Mahesh for his second part. Thank you. Thank you, Teresa. That was an excellent presentation on the insights into lipids quality. So hello everyone. Welcome again. I'm very excited to move into the next segment of the presentation focusing on the process development and scale up of LNPS and bringing them into clinical and commercial manufacturing. Would like to thank you all for being here to hear what we have to say today here. So here is what a brief outline of my presentation going to be. I'll begin with an overview of unit operations involved in manufacturing of Lmps, briefly touch upon the capabilities of process development and optimization with few case studies. Then I'll move into scale up and GMP tech transfer and then I'll wrap up with capabilities on analytical development of lmps at the end of our presentation. Hopefully will address the overall goal of how we can collaborate with you to help develop your LNP products into commercial drugs involving breakthrough therapies with the common objective of saving patient lives and are improving their quality. So going into a little bit of introduction of this topic. So here are some schematics that you see of some important lipid based drug delivery systems. You see lnps on the left here, then liposomes and lipoplexes. And these are all examples of drug delivery vehicles. Not to say that they naturally occur in this vibrant colors, however you can visualize them as nano sized balls made of fat. That carry the desired payload in them. Now recently COVID vaccines have demonstrated beyond doubt the ability of the LNPS to deliver nucleic acids and are currently touted as the most popular non viral gene delivery system now going into structural nuances of these nano formulations. The lipid nano particle here. It consists of a lipid shell consisting of an internal core, typically non aqueous, composed of several reverse micelles that encapsulate the oligonucleotides like SI RNA mRNA, G RNA plasma DNA. As you see, they typically do not show a lipid bilayer. In contrast liposomes they contain one or more lipid bilayers. They further classified by size and lamellarity in the form of small, large or multi uni lamellar vesicles. They contain kind of an aqueous core. Now they can kind of encapsulate. Hydrophobic molecules either within the lipid bilayers or can encapsulate hydrophilic small molecules within the APS core, moving on to lipoplexes. Lipoplexes are defined as the complexes of plasmid DNA and cationic lipids. They are multi lamellar liquid crystals consisting of hydrated DNA layers. Alternating with cationic lipid bilayers, they do exhibit electrostatic complexation between the cationic lipids and the phosphate groups of Dna's. So giving an introduction here into the capabilities of the site here. So the site here is set up to perform phase appropriate drug development activities. At any stage of the drug life cycle from preclinical through commercial, we have clients that we collaborate with whose programs are at various stages of this life cycle. There are some clients that we collaborate with at preclinical stage with a complete development scope, process development, analytical development. Where the formulation is kind of almost set here and we are able to on board them with their very preliminary or an initial process could be at sub mill volumes and then we are able to scale it up all the way say from mills to multiple liters in the pond space. Now there are kind of other client projects where we are able to on board them. We are tech transfer directly into our GMP space without any development scope involved there. Now there are also few projects where we've kind of onboard them early on in the phase and are able to optimize their formulation and kind of are able to scale them and move them into later phases. Here what do I the thing that I want to mention here is that. The site offers a complete end to end solutions including aseptic fill finish, packaging and inspection, QC and micro testing, CMC support, project and supply chain management. So you can see we have kind of all the capabilities under the belt here. So we will move into the next poll question in the series. The question is what is your current drug product development stage, a preclinical B phase, 1C phase two or 3D commercial. I will just give you a little bit time to kind of reflect and vote on this question. Wow, it is a complete winner on preclinical here. Thank you again. So moving into kind of the meat of my presentation here. So for simplicity, the manufacturing steps or unit operations in LNP or liposomal products can be broadly categorized into four separate unit operations. Now the first step here is the nano particle formation. Which I consider as the heart of the LNP process. The schematic here shows an example of inline mixing process of API based aqueous stream and lipids based ethanolic streams where they are kind of mixing through a suitably designed mixer. The stream is then diluted with a dilution buffer which is altogether collected. As a pre bulk LNP material where it basically consists your lipid nanoparticles here at this stage. Now I do want to mention in addition to in line mixing the site also here has experience in the development of ethanolic injection and impingement jet mixing based processes. Altogether these are the commonly available processes that can be scaled up seamlessly. But however we are able to work with the process of yours as well in terms of the nano particle formation step here at the site. Moving on to the second step which involves purification or a concentration adjustment step of the LNP pre bulk material which kind of needs to be freed from ethanol. From salts and small API molecules as well which leads to pure LMP material. Now this purification is accomplished through the tangential flow filtration consisting of ultra filtration and dia filtration steps. The Dia filtration step involves buffer exchange from a typical citrate based acidic buffer to a neutral buffer. The Ultra filtration step involves concentrating the material to target concentration level. The material is then diluted with a cryo protectant buffer. The third step involves filtration and clarification sterile filtration I should say, which ensures that the material is sterile and free of bacterial microorganisms like other biological products. Which are quite sensitive. Terminal sterilization is not possible given that they are very harsh to these products. Here the material is pushed through a point to Micron membrane to obtain sterile bulk material. Now the final step or the unit operation involves here is fill and finish where the product is filled into a pre selected bio size with a predetermined fill volume. Based on the dosage and administration requirements in the clinic, the vials are then right away taken into cold storage, either minus 20 C or minus ADC or they can actually be even shipped for lyophilization AT28. Let's now dive deeper into some of these unit OPS. Now. As stated in the previous slide, the LMP mixing step right is the heart of the process. Which leads to production of nano particles with defined characteristics. We have helped several clients in their journey in developing this step from a very clean slate. The site here possesses the necessary experience and skill set to help design your LMP, mixing skid and the overall process we typically deploy turbulent mixing. Technologies for ease of scale up, but not to mention we will also partner with you on a skid of your own choice as well. Now think of this whole mixing design skid as a one big toolkit with various parts to choose from. So first is your pump technology. Now the site here has experience with various kinds of pumps for the scale up. Could be a crystaltic. Air pumps, HPLC injection pumps and diaphragm based pumps. Now what is critical here is the ability to deliver accurate, precise and stable flow rates over a long duration with minimal drift and minimal shear impact to the lmps. And what is also important is the ability to modulate the flow rates appropriately with the room to upscale when needed as the clinical stage progresses. The custom versus off the shelf mixers. These are very critical and precision design tools that can help generate and tune your particles with consistent size and PDI and encapsulation. Now we can help design your mixers with off the shelf parts or work with a choice mixer of yours that's highly customized as well. Typically these include Ty Cross mixers, dimensions with very precise ratios of inlet and outlet diameters. Then there are several critical process parameters that we define. At this stage we can help determine or vary or even monitor the concentrations of your input materials API and lipids, dilution buffers, flow rates of all of these streams. With precise floor pressures and temperatures now moving in to the process development and optimization now the slide here describes the several process parameters which give the ability to tune the physiochemical characteristics of the nano particles. A stable and a robust process which delivers a consistent product. Quality is what is required here. Using design of experiment methodologies, we are able to develop a generalized strategy to optimize the physico chemical characteristics of the lipid nanoparticles with the ability to vary concentrations of lipid to API ratios, flow rates. Of both the API and the Ethanolic streams, the injection ratios of the aqueous over ethanolic streams, we can kind of design this doe to two to three levels such as low, medium or high and we can kind of create a design space to understand and generate particles of desired characteristics size. Pdi encapsulation and the lipid to API concentration are typically relied on as CQS here. We will further elaborate in the next few slides on this capability with some example case studies. We have also helped clients to further optimize their formulation by changing buffer type, pH, Osmo and an appropriate cryo protectant buffer. The robustness of these formulations as well as the process can be rapidly assessed via specified stability studies can be stress accelerated normal to verify long term stability and ensure that the formulation as well as the process is robust to deliver a stable product. Now here I am going to present a case study have in a very semi quant manner in a very generic manner involving the process development of a RNA based LNP generated via turbulent mixing. The intent here is to just showcase our process development capabilities. Now this data was generated as part of a DOE study where lipid concentration. Flow rate which is Fr designated here as Fr and injection ratios, the IR and the encapsulation efficiencies as percent EE. Now where we modified the lipid concentration, flow rates and injection ratios and kind of studied their impact on the size and PDI and encapsulation. So the first plot here shows. How the LNP size is impacted by varying lipid concentration and we are also kind of varying flow rates as well as injection ratios. As far as the DOE now the flow rates are shown by the color-coded legends. The high flow rate is shown as a green circle versus low flow rate as Red Square here. Overall what we see here is that with increase in lipid concentration and coupled with flow rates. Especially the the low flow rates here you see that the particle size is kind of increasing, but the spread of size is also becoming tighter. With the increase in lipid concentration, you see the spread as minimized here now with respect to flow rate. What we saw here was there was an inverse trend where high flow rates shown by these green circles, they led to particles of lower size relative to low flow rates shown as red legend, which led to kind of higher size. Now in scenarios where the lipid concentration, the flow rates were similar as you see here they are kind of lined up. Now what is it that is giving the ability to attenuate particles of either higher sizes or lower sizes and that is the injection ratio. So higher injection ratio of the aqueous to ethanol extremes kind of led to particles of higher size. And what you see here also where flow rates and the lipid concentrations were similar, high injection ratio led to particles of higher size versus low. Injection ratios. Now moving on to the second plot here. This plot here shows the impact on the PDI of the lnps. The parameters here are same as what we discussed here in the previous plot. We have the lipid concentration which we are varying here on the X scale. We also have dissimilar flow rates low, mid and high. All of them shown in their color legends here low as in Red Square and high flow rate as in green circle. Now what do you see here is that in general the PDI overall decreased with increase in lipid concentration. So you see here these are the PDI and a kind of decreasing going as how the lipid concentration progresses. Now what you also see here is. This spread in the lipid concentration is kind of minimized while you sorry, the spread in the PDI is minimized as you increase the lipid concentration. The similar impact of injection ratios was seen here as well where if you have a scenario where you had similar flow rates shown by low flow rates and similar lipid concentrations, higher injection ratio led to. Higher PDI versus lower injection ratio which is aqueous to ethanol extremes led to lower PDI. Similarly you see this with on the green circles where if you had similar high flow rates, similar lipid concentrations, it is the injection ratios that impacted the PDI. So let us move on to 1/3 set here. Now this data set shows the optimization of LNP encapsulation. Versus lipid concentration. Again as far as the DOE, we similarly modified flow rates as shown in the previous plots. We also kind of modified injection ratios. Now in general what we saw here is that it increase in lipid concentration coupled with low flow rates shown by these red squares. This led to higher percent EE. Now again in data sets where. The lipid concentration and flow rates were similar as shown by the red legends. Here the IR had the ability to impact the percent EE similarly as shown in green legends with higher flow rates. Again, low IR led to high EE while the high injection ratio resulted in lower EE. What I want to alert here is that the trends that you saw here in percent EE, either PDI or the size were kind of very specific to the formulation and the mixing skid that we had developed for the process. And it is very likely that these trends could change with a different formulation or mixing skid. So in general, what I want to summarize here is that we were able to optimize. The LMP size, the PDI and percent EE by this doe involving all of these three parameters here. So let's move on to our next segment, which essentially covers the LMP downstream processing. Now these downstream processing steps ensure that the lmps are homogeneous. And stable during storage and they are free of residual contaminants. So in terms of these processing steps, we deploy extrusion, we deploy tangential flow filtration now on the extrusion. This step reduces particle size and generates uniform particle size distributions. Extrusion involves essentially pushing the bulk lnps via pressure through membranes. Whose pores can range from 80 to hundreds of nanometers and you can probably have one pass of this pore shot to multiple passes. Now this step is generally performed with liposomal based ethnolic injection processes and not typically used on LNP based processes. I do have to say that in terms of the capabilities, the site here has the ability. Process volumes ranging from melts to hundreds of liters and handle pressures up to 800 psi or more in order to remove bigger particles and or clean up the tail end of the particle size distribution which is the D90 and D99 typically and this can help decrease the PDI essentially. Now moving on to the tangential flow filtration. This is a schematic that shows an example of the setup of tangential flow filtration. So we have the ability at the site here to optimize your TFF processes. We have capabilities to perform the ultra filtration and dia filtration steps ranging from 50 mills all the way to hundreds of liters. We have pumps that are low pulsating diaphragm based that are able to deliver flow rates up to about 30 liters per minute in the PD space and also have. Larger flow flow rate capability in the GMP space. In the P space, we have scaled up from 26 centimeter square membranes all the way up to 5.7 meter square membrane surface area. Using appropriate membrane and material type in kind of both formats, hollow fiber membranes as well as cassettes, we are able to regulate the cross flow rates, the trans membrane pressures. Shown as TMP and the shear so that we can minimize the overall TFF time and improve the permeate flux while maintaining the product quality without traveling the membranes. The cryo protectant buffer can be introduced during DF or during the dilution step after UF and PREFILTRATION. So moving into the next step. Here the clarification of the filtration. Of given the requirement of sterility assurance and sterile filtration kind of being the means here, this is a very critical step. The criticality of this step is further more significant in the context of LMP and liposome products. In order to mitigate any filtration challenges and thoroughly understand, we recommend to characterize the tail end of the Z average distribution in the form of D90 and D99 characteristics. Again the membrane type, the construction, the double layer of the membrane can help with this. During the clarification stage to remove any debris or tail end of the Z average in the form of bigger particles, we scale up the filters appropriately based on the volumes in hand. We are also mindful of filtration losses for small scale batches when planning product delivery, we are able to monitor pressures. Volumes or weights during the operations and determine flux and throughput filter validation again which is a requirement here at the site. We work with multiple vendors to able to kind of execute this step here. So in summary, we have a wide range of capabilities and experience for producing. Small scale as well as scale up batches in terms of both the scale sets and as well as tools. So moving into the next segment, the process scale up. In GMP tech transfer, the process developed here is transferred as is from the PD to GMP space, the mixer ID, the flow rates, the tubings diameter, length of the tubing, they are all typically fixed now. The scale up is performed as a factor of time. Where the process is allowed to run longer till the RNA or lipid stock solutions are consumed up. We recommend to evaluate the material stability from the standpoint of mixing and processing durations. Since the product is sitting at room temperature in low pH buffer, we also help out in the selection of tank sizes, consumables and scaling up of your filter surface area for procurement we try to engage. Our supply chain early on, so the impact on lead times is mitigated from a timeline standpoint, single use product contact surface areas are preferred. So the requirement of cleaning methods and calls are minimized. We would recommend that there is not more than ten time jump from a PD to GMP scale up. So the dynamics and process parameters are kind of representative going from PD to GMP. Lines are supported on various scalar batches and planned early on to help coordinate with the delivery of material for the plant studies. The talks batches in PD are de risked in the open PD space by performing various mitigation activities. The filling is typically performed biosafety cabinet. Here we collaborate and coordinate early on with the process engineers in our tech support department. With drafting up of batch records and SOP documents and other documentation required for GMP, the site does have a filter validation as I said previously from austerity assurance standpoint. Now in terms of process controls, we are able to monitor and determine your flow, temperature and pressures. We help design the in process testing at. Your critical operation steps and make sure if they need to be gated and make sure we recommend the process whole times based on the dynamics of how the process runs in the GMP. If there is shared equipment between products, we also help recommend cleaning requirements and validation early on South. As a summary here we support in a complete. Planning of your material procurement, procurement as well as a scale up early on to ensure a very successful tech transfer. Now moving on to the last segment of my presentation, which is the analytical development. Now the site here has extensive experience in terms of depth and in terms of breadth in terms of skill set or experience and method development as well as executing method calls on your methods. Of course, on the critical quality attributes like particle size, we have capabilities, API encapsulation efficiency, which is typically performed via ribogreen. For RNAs, the API concentration is also performed via orthogonal assays in addition to ribogreen such as iprp or SEC. The API purity is. Performed through fragment analyzer, sometimes the smaller APIs such as say RNA, we are also able to help perform iprp lipid content and impurities are evaluated typically through UPLC, CAD or USD based methods. The residual solvents are determined typically which is the ethanol. Here via GC fit. The Physico chemical characterization is performed to zero potential pH, osmo and particulate matter. Now the raw material ID testing we recommend in the early phases typically is FTIR or UPLC based methods and are able to ensure that we are pulling right materials into our manufacturing rooms. We do also require that there is micro testing on these materials, but otherwise this micro testing could also be performed here at the site. Now in terms of micro testing we have a. Complete department that is able to perform sterility, bio burden and low endotoxin testing on your products. And now in summary, I have to say that the site has extensive experience in analytical development and GMP method calls. So as a summary on my presentation, I have to say the site has successfully manufactured several clinical LNP products. We also have extensive experience in process and analytical development including optimization and troubleshooting and we have applied them successfully on several projects as well. We have successfully scaled up and GMP Tech transferred many, many projects at the site. So we can truly be your partner in terms of an integrated offering like summarized by Teresa on the lipids, we are also able to. Procure and supply and manufacture your mrnas and then we are able to use these materials to develop your process formulation manufacturing all the way up to fill and finish. So in summary, you have seen Teresa talk about the critical quality, quality considerations for your lipids. I have kind of. Given a deep dive on to help bringing your LNP development all the way to commercial manufacturing. So here are the three key takeaways from today's presentation. Combined with mine and theoriesis, the lipids quality and characterization of impurities are crucial for the successful development of LNPS for mRNA delivery. Second, the expertise and capabilities in process and analytical development are essential to ensure a smooth and a successful GMP tech transfer. And as a summary, again we deliver end to end services providing unique and integrated solutions and offerings all the way from materials excepients to final direct product. So with this, I kind of wanted to thank you for your participation and your attention today. Thank you very much, Mahesh. And thanks again also to Teresa for the great presentation. We have several questions that have been coming through from the audience throughout the session. Just before I jump in and and focusing on the questions and and give back to to the speakers to to answer them. I just wanted to remind that you can still submit your questions using the Q&A widget and if we don't manage terms of times to cover all the questions, we will reply to you. We'll respond later on via e-mail. So keep the questions coming. All right. So let's just jump in. We have combined some of the questions and maybe we can start with Teresa on on one that is focusing more on the key excipients that are unnecessary to build the LMP formulation. So Teresa, can you cover again, I think you had also a slide during your, your part of the presentation on what are the key excipients to build LMP formulation, sure, yeah. So you can see it now here. So we need, due to the various function, different excipients and thus different lipids in order to manufacture lipid nanoparticles, and you can see them listed here. So another question on focusing on the lipids for you there is. So we are still doing preclinical stage formulation development. So the question will be is, is it necessary, should we already consider GMP lipids at this stage? Yeah, it's a very interesting question that I don't hear it for the first time and I think today both me and Mahesh have highlighted. How decisive the impact of the quality and the chosen raw materials can be during the development journey. And so we recommend to keep an eye on that as early as possible. Because if you choose, for instance GMP lipids that from preclinical you can be ensured that if you change them later on throughout your development process, you will not have significant changes in. Your drug product profile or even your process. So therefore we recommend to consider quality already when we talk about preclinical stages. Great thanks Teresa. Also checking time, another one for you maybe do you provide regulatory support for your lipids? So let me differentiate here between portfolio and the custom service that we offer. So for commercial grade portfolio limits, we do usually have DMS filed already. So then customers can get access via a letter of access to the filings. For custom manufacturing, we provide service for the product that is then in in the development target for every stage required by the customer so we can support. Early stage filing, up to commercials filing. So the general answer is yes. Thank you very much. Maybe now we can move to to Mahesh. We have several questions that you can probably cover. Let's start with this one. I think you covered this already during the presentation, but maybe could you explain again? The LMP formation to fill and finish process, sure. You can also show, I don't know if you want to show anything on this screen. Would you be able to show the slide #22? Yeah. So I'll just show all of these unit operations all together in one slide. So the question is, can you repeat the question again, Francesca? If you can explain basically again let's say the process from LNP formation to fill and finish, so summarize, right. So I mean it's typically 4 unit operations that the LNP processes does have. One is first is your nanoparticle formation which basically involves lipids in ethnolic streams and AP and AP Streams which are kind of pushed through a suitably designed mixer. And they collected as a pre bulk material. If there is a dilution buffer sometimes as well then this LNP pre bulk material is kind of purified through tangential flow filtration process, through a buffer exchange as a DIA filtration step and then through ultra filtration steps. It's kind of got into a circuit certain target concentration. So here. Essentially we are getting a pure LNP material which is free of ethanol. It is free of salts and maybe sometimes small AP molecules. And then we take this and add either the cryo protectant buffer during this step or either during the DF step and kind of perform a sterile filtration on this bulk material and then go up to fill finish all the way. Hopefully this summarizes. And and kind of answers the question. Thank you. And also as a reminder the webinar will be available on the month. So you can also go back to the presentation in case you want to to check again on on this specific topic. All right. Another question for you Mahesh, for clinical trial, what is the scale that we have to prepare for? One liter, 10 liter more the clinical trial, we can actually target any liter size that we want like all of this target would basically be involved in scale of batches that we develop in terms of the process. So I did mention that we typically recommend not to do a 10X jump. So if you say want to target a 10 liter as part of your clinical. Then we want to make sure that the process is developed at least up to couple liters so that the there is a seamless transfer here, but we can even develop up to hundreds of liters. But we want to make sure that we have appropriate skid that kind of supports the flow rates required and ensure again that the particle formation step that it the time it takes to form the particles is kind of conceived consciously so that it is not very, very long. So to answer the question, we have capabilities to hundreds of liters to on the large end and even personalized product to liters one liter to multiple liters. Great. And that now moving from the larger scale, one question on, on the smaller scale, so can you share on what kind of instruments do you use for LMP manufacturing at small scale? So at small scale, again the instrumentation is going to be similar as to what they're going to be used in the large scale. Again, the pump technologies, the mixers, they're all going to be same. If you have used on the small scale say gear pumps or maybe syringe pumps as part of your IGM process, we will restrict to that. We will use mixers. And the same mixer, the similar pumps, everything would be kind of used in the scale up and the GMP tech transfer more or less great. I think we are at time unfortunately we have many other questions but as I said we will we will tackle this and reply to you individually via e-mail. So thanks again Mahesh and Teresa for today's presentation. Thanks to the audience and everybody for joining us. And just as a side note, if you want to register to future webinars or access our webinar library, you can visit our website. Have a great day ahead. Thank you all. Thank you. Bye. _1732521335431

 

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