Discover Many Receipt Formats for Research and Development Made Easy

Many receipt format for Research and Development

good morning everybody my name is rhonda henshaw powell i'll be your moderator for today where just a couple of housekeeping requests before we get started please remain on mute um this can be particularly distracting for both of our speakers today and for the audience as well and also we have our series of panelists available online some of our acd um support and pas experts waiting in the wings to answer any questions that you have during the presentation so as you're listening to today's speakers please do pop your questions in the questions box and our team will be addressing them as we go through we have two excellent speakers today shawna levinson and alpana kumari both members of the pas and support teams and they're going to be presenting uh the clinical assay development process using acd's ish assay services so with that i'd like to hang over to our first speaker shauna levinson thank you very much hello everybody uh hope you are all staying safe and healthy we are excited to have you here today and present to you about our expertise in rna-ish and how this can be applicable to your clinical essay development needs again my name is shawna levinson and i am a senior research associate here at acb and part of the pharma asset services team i will be going over the rna scope technology as well as the cdx development and clinical assay workflow i will then hand it off to my colleague dr rapana kumari to go over examples of rna scope use in the clinic [Music] i'd like to start with a snapshot of biotechne as a whole final technique combines a wide variety of brands companies and solutions that are available to help with your research so what is rna scope and how is it relevant to space spatial transcriptomics traditional transcript analyses have relied on bulk tissue analysis such as pcr which provide an average level of gene expression in a sample but loses the spatial and single cell information this is akin to a root smoothie in which you can only guess at what fruit it contains the advent of single cell transcriptonic analyses such as single cell rna sequencing have now been able to provide molecule information molecular information at the single cell level and to come back to the fruit analogy can now provide you the menu and tell you what fruit and how much of each fruit went into the smoothie however these techniques still don't provide information on how those cells are organized in the tissue context this is where spatial analyses come into play single cell multiplexing spatial analyses that detect rna can provide transcriptonic information with spatial organizations while retaining single cell resolution so now instead of an average blended smoothie with mystery ingredients you can see all the fruit and how it is spatially arranged the growth and adoption of rna scope technology is best exemplified by the number of peer-reviewed publications we had our first publication in 2011 and since then there have been over 2700 peer-reviewed papers published using the rna scope technology in numerous journals including many top tier ones the arnesco technology is highly relevant across multiple fields of research with over 40 percent of the publications in cancer research followed by neuroscience and infectious diseases furthermore the incorporation of the rna scope technology into single cell rna sequencing workflows is demonstrated through the rapid growth of publications using the assay to confirm and spatially map single cell rna sequencing data in the tissue context with over 80 of these single cell papers in high impact journals including nature cell and science as well as top journals in specialty areas such as pathology oncology neurology the rna scope technology is an ideal spatial analysis solution to interrogate complex tissues it is highly specific and sensitive method to detect rna biomarkers and cells and tissues with morphological contexts at a single cell level it consists of three parts a unique target probe that acd designs against your sequence of interest a signal amplification system that generates a high sigma multinoise ratio and lastly visualization of single rna molecules as dots the assay allows for spatial mapping of mrna long-encoding rna splice variants point mutations and highly homologous and short sequences in cells and intact tissues of which can be visualized with either fluorescent or chromogenic labels the essays can be performed on a wide variety of sample types including ffbe tissues fresh frozen or fixed frozen tissues pbmcs and cultured cells diving slightly deeper into how the technology works the first key feature of the rn scope technology is the probe sign we depict the eleven nucleotide target specific probes and z's to emphasize the fact that they have two regions linked by a spacer the bottom of the z complements the and hybridizes to the target transcript the top of the c is the base for the amplification structure when the two z's in its easy pair hybridize in tandem to the target sequence it creates a unique binding site upon which a pre-employer can bind and the amplification tree can be built a standard ornascope probe for a target sequence of a thousand bases or more will consist of 20 zz pairs pulled together and designed to hybridize in tandem along the target region this allows for a tremendous amount of amplification and signal potential however only a few zz pairs are needed to bind to the target rna sequence in order to generate enough signal for molecular detection here we walk through how signal is amplified with the rna scope assay the two key features of our technology are the promising as we just described and the signal amplification system once the zz pair is bound and tandem to the target a preamplifier uh can bind the opposite end of the pair after which the amplifiers bind to the pre-providers followed by which the chromogenically or fluorescently labeled oligos bind to the tree this amplification system allows for visualization of single rna transcripts and hence very low levels of target can be detected on the other hand if only one z of a pair binds down specifically to a sequence the preamplifier will not be able to hybridize and the amplification tree will not form as a result no signal will be detected the probe design and amplification system at strategies provide for high sensitivity and specificity of detection single molecule detection is what we focus on and what is possible with the rna-scope assay this is illustrated in an experiment that was published in the first publication demonstrating rna scope yard escobace one plate of heloceph culture was run using the rna scope assay probe for her2 and the other was lysed and then the solution was broke for her too with the rna scope experiment on the left you can count her two mrna as the reinforcement dots which averages about 14 dots per cell or 14 or two rna copies per cell on the right a standard curve was used to determine the average for two rna copies per cell and the lysate which averaged about 17 copies per cell so the two assays give very similar rna copy readouts uh the point being here that when we see a dot using rna scope it translates to a molecule of transcript here is an example of rna scope being used to tease out heterogeneity and gene expression within the tumor this is a duplex chromogenic staining for pdl1 and lag three which are two common targets in amino oncology it was found that in this tumor pdl1 is primarily expressed in the tumor compartment whereas like three is primarily expressed in this dermal compartment this information is lost in the whole sport the whole core analysis the rna scope technology includes three unique assays to detect rna in the tissue context at single molecule sensitivity with single cell resolution this means that a single dot represents a single rna molecule or target these three assays are rnascope vascope and micrornas the rna scope assay is applicable for mrna or non-coding rna greater than 300 nucleotides in length and is available in singleplex or duplex chromogenic or as multiplex fluorescent assay which can detect up to four targets simultaneously face scope is detected for the i mean design for the detection of exon junctions splice variants point mutations and highly homologous or short sequences between 50 and 300 nucleotides in length phase scope is available as a single plex or duplex chromogenic assay my microrna scope is designed for the detection of antisense oligos micrornas sirnas and other rna sequences between 17 and 15 50 nucleotides in length and is available in chromogenic parting now i'd like to give you a brief overview of acd's pharma asset services pas is a team of 20 plus highly trained specialist scientists and pathologists that are experts in rna scope performance and analysis that will ensure delivery of the best data possible we provide services for 20 of the top 20 requirement companies in the world in general we deliver greater than 15 000 stain slides per year to customers in more than 20 countries as shown here we can help with acquisition of tissue samples through our trusted chain of tissue providers and qualification of these for the arnoscope assays we also work with third-party cros and can easily manage the embedding and sectioning of the samples in your studies we perform the rna scope base scope and and microrna scope assays which are routinely provided through our service panel provide one quantitative assessment and put all this together into a report which includes our findings as well as the materials and methods of the study we also wanted to share with you something about how we manage quality systems and security for the data generated through pas sops govern the pas workflow including staining procedures sample tracking data management and data pc each essay run includes hela cell technical controls the hila positive and negative control slides must pass strict qc requirements in every assay run any runs that fail the run suitable suitability requirements will be repeated the data that we generate here is shared through a secure enterprise file sharing system called ignite access to this data is highly restricted the pharma assay services team can help you with drug discovery through the phases from target discovery to development we can help with target screening efficacy screening dmpk studies safety and toxicology studies as well as performing early feasibility assessment for development in phase one clinical trials here we have pas the pascdx and clinical assay development workflow for massey development and precision testing the pathologist guided scoring and analysis the written sop and reporting to assay transfer and cross validation this slide illustrates a sw or analytical plan and essay development here on the left you see a pharmacy services so w template asset development would include choosing an assay platform it might include optimization steps for sample pre-treatment and probe optimization [Music] and acceptance criteria our validation parameters include specificity and sensitivity repeatability and precision dynamic range robustness and stability essay precision testing might include inter-instrument and intraday reproducibility as well as same-day repeatability bas also uses pathologist guided scoring criteria for sample analysis as seen here a pathologist has appropriately annotated the different tumor and tumor associated stromal compartments for the identification of heterogeneous expression patterns these histopathology notes can also include things like tumor load and visual age scoring the pathologist is also available to discuss and review data we also wanted to share with you how our lab manager and project manager work very closely with the clinical specimen collection site as well as the central data management lab to work through data transfer specifications and agreements our project managers meet on a regular basis with the sponsors and data management sites for smooth project and data flow lastly as you see at the bottom there we have a streamlined process for batch processing of samples as they come in and we work with our sponsors to provide these to report data back with minimal timelines this slide demonstrates an example of a batch workflow for 1 to 10 samples per batch and depending on the assay on day one we would receive the samples and begin qc staining day two would include post-processing and review of the qc slides day three would include marker staining and day four post processing and slide scanning on day five there would be side scanning scoring and analysis and day six would include reporting data reporting data qc and project delivery pas utilizes labvantage which is a widely used limbs in the industry for auditable and efficient sample tracking throughout studies for gclp guidelines and to ensure data sample and sample security the pharmacy services group is here to provide you with data and to form any of the rna scope assets for you the pas group has profound expert expertise with over 15 000 slides delivered per year access to 14 automated staining systems from leica and ventana digital slide scanners for both bright film field and fluorescence images halo software for quantitative image analysis board certified pathologists are on staff and a tissue bank with qualified human normal and tumor tissues available for immediate screening once your study is complete you will receive a report containing representative images materials and methods summarize data analysis and the study summary and conclusions you will also receive full slide digital scans the same size and any unused material acd has established protocols for clinical assay transfer to clia certified certified labs for respective use invalidation cross validation we provide three way conference calls between acd and sponsor and the clia lab to go over things such as essay and protocol transfer and verification and cross-validation and pathologist scoring criteria we can also provide an on-site visit with one of our field application scientists to assist in the transfer process and now i will uh trans i will hand over the presentation to dr alpana kamari to go over examples of rna scope use in the clinic hello everyone uh can you hear me yes we can thank you el pala hi uh my name is alfona kumari and i am a member of pharma asset services at acd and in the next few slides i will show a few cases case studies illustrating that rna scope has been proven to be effective means of detecting a biomarker that would be predictive of patient response to cancer therapeutics first of all i would like to shed some light on the features of rna food technology that makes it suitable for diagnostic tests rna scope works best with sspe tissues which can be fresh or archival tissues in addition to fse rna scope works equally well with frozen samples and the flexibility in the sample type is certainly helpful for clinical studies the rn expression data can be visualized with morphological context and its sensitivity is at the level of single rna molecule which is much higher than isp the data analysis also allows flexibility as the scoring can be done visually by a pathologist or it can be quantitative quantitatively done using a software finally it is performed on automated instruments using in ensuring that it will be reproducible between batches this is a chart comparing irony scope with other available essays and as shown here rna scope is the only platform that provides most sensitive and specific data in morphological context and it is flexible in terms of tissue requirements as mentioned earlier it can be performed on almost any type of sample be it ffe fresh frozen fixed frozen tissues or cell lines furthermore specific rna scope probes can be designed for any target or biomarker it can be performed as a single flex using one probe at a time or it can be multiplexed to visualize multiple targets in specific cell types and finally the quantitative data analysis will provide cell by cell data with single molecule sensitivity this is a piece of data from clinical cancer research published in 2014 showing call 11 a1 protein and rna expression in primary metastatic and recurrent serious ovarian cancer tissue the authors saw good correlation between protein and rna during cancer progression rna scope showed better dynamic range of expression than isp and rna provided higher resolution than protein as we can see here the top panel is showing isp and the bottom panel is rna scope and it is clearly visible that rna scope is showing a better dynamic range as compared to isp therefore they determined that rna's rna serves as a better essay for call 11a1 as a biomarker for serious ovarian cancer we do have a validated protocol for immuno-oncology checkpoints target profiling and this slide is demonstrating the importance of tumor heterogeneity these are the snapshot images showing expression profile of various io targets in two different non-small cell lung carcinoma tumors clearly there are differences in expression pattern of these io markers between these two tumors for example lag 3 is showing expression in both tumor and stroma in first tumor however it is only expressed in stroma in the second tumor and similarly there are differences in the expression pattern of pdl one thin three and pdl2 between these two tumors one important observation is that sym3 is expressed in almost mutually exclusive pattern here excuse me with pdl1 but these are co expressed in the second tumor in in in some of the cells and this coexpression analysis of immune checkpoint marker at the single cell level could potentially aid in understanding its resistance mechanism of checkpoint inhibitors as well as it can bring insight into combinatorial therapies one of the most widely used diagnostic application for rna scope is the detection of hpv in various tissue types including head and neck cancer cervical dysplasia and cervical cancer stages can be used in the clinical decision making process to select patients for less aggressive non-surgical treatment and we all know that p16 is a well established surrogate marker for hpv status however e16 can be induced by other pathways which are independent of hpv and this kva led to interest in more specific assays for hpv detection among those assays are dna and rna scope in c2 hybridization for hpv and these essays have been evaluated and there have been tremendous amount of work to evaluate and compare specificity and sensitivity of various assays for detection of hpv as shown here rna scope is the highly sensitive and specific essay that can provide direct evidence of hpv activity in this slide on the right side i'm showing a kaplan-mirror curve which which shows that the p-16 positive but hpv negative patients they have significantly reduced five-year survival for oropharyngeal cancer as compared to the patient which were p16 positive and hpv positive and this p16 faulty but hpv negative was almost equivalent to p16 negative now since p16 isc may provide a false positive report for hpv status reliance on p16 isc is quite risky on the other hand acv rna scope testing is more specific for determination of hpv status and it can provide favorable prognosis so for the development of rna spoof based diagnostic tests fed has partnered with like a biosystem this partnership leverages the combination of acd's rna scope infidel hybridization technologies with lycast bond clinical advanced staining instruments uh nasa has two types of automated histology system the first one is bond rx and the second is bond three these are basically identical systems however bond rx is mainly used for research grade tests such as biomarker validation pre-clinical studies asset design and feasibility and prototype testing and development on the other hand bond 3 is fda approved system which is developed under design control and the assets performed on bond 3 can be fully validated and registered with the fda based on being performed on this fe approved instrument the first clinical diagnostic test program that we worked on was with merrimack pharmaceuticals for their anti-herb b3 antibody therapeutic called mm121 they had determined that an rg1 which is also known as hergeline was the biomarker that was correlating with patient response to treatment with mm121 in combination with standard chemotherapy peridoline is a secreted protein so ifc was not an option and they would not get enough material from a needle biopsy to perform qrt pcr where whereas it was enough for detection with rna scope and as we can see here these dots are showing are the uh peripheral message that that was detected in the small biopsy and this is a negative uh control probe this is a positive control probe and uh the the clear background with negative control and the good standing with positive control is giving us the confidence that the the signal that we are seeing with herogalin is the real signal so finally uh they partnered with leica biosystems and utilized rna scope essay for combinat combination diagnostic tests i'm sorry for companion diagnostic test development and for the selection of patients in phase 2 clinical trials this is the data from their phase 1 study and this is showing progression free survival in patients treated with mm121 it shows that breast cancer ovarian cancer and lung cancer patients with high hearing healing responded much better when they were treated with mm121 in combination with standard of care therapy as compared to standard of care therapy alone for example here these blue lines these are the combination therapy with the patients having a high herugulin level and the black lines are the ones with the single chemotherapy and it clearly shows better progression um free survival in these patients this is a screenshot um of a poster presented by merrymac and this is not a very uh clear uh poster but the the message that i want to send here is that they presented this poster uh with their academy collaborator at astro in 2019 and uh they reported that uh their results of the phase two clinical uh trial studies that that mm121 was used uh as a combination therapy and they performed it on almost 500 patients which came from 87 clinical sites on three different continents and the performance of iron scope was highly robust and reproducible on this large number of patients another example where rna scope has proven to be most effective for biomarker assessment and development of diagnostic tests is for receptor tyrosine kinase fgfr 1 2 and 3 for which bayer has a small molecule tyrosine kinase inhibitor rna scope was shown to be superior in terms of both specificity and sensitivity against isp as well as nanostring assets and originally in 2014 rna scope was proven to be a better predictor of fdfr tyrosine kinase inhibitor sensitivity as compared to gene amplification and it was also shown to be more specific than iec based on these results they chose our mrna essay as a basis of enrollment of patients in their phase one clinical trial and the positive results in that phase one clinical trial led them to partner with leica biosystems to have fgfr companion diagnostic as the development based on the rnesco technology rna scope was then used for patient selection in a pivotal phase 2 and phase 3 clinical trials shown here is an example of a patient who was highly responsive as you can see here uh between the baseline and the 11 day week of treatment this patient was highly responsive to the bare therapeutic tyrosine kinase inhibitor and this was kind of this was negative for sdfr amplification however the patient did show high level expression of fcfr messes into your biopsy as you can see here for further illustration this is again a snapshot of asper poster describing the use of sdf rna expression as the basis of patient selection for phase one trial and this trial involved 860 patients on which they they ran fcfr 1 2 and 3 tests again illustrating performance characteristics and high level of reproducibility and robustness of a clinical rna in c2 hybridization assay this is a piece of data uh from leap therapeutics that they presented at sixty in 2019 and they showed that dkk1 expression correlated very well with a response to the treatment and here the top panel is showing the biopsy patient biopsy which showed high expression of dk1 with its score of 163 and the bottom panel is showing a tumor biopsy with low level of dk one with his h score of seven and on the right side is the kaplan major curve which shows the the better survival of the patient with high tk1 level as compared to the one with low dkk one level and therefore they are going to use rna scopes as their patient selection test in their upcoming clinical trial where patients will be selected based on having certain threshold of bkk1 expression this is another example of use of rna scopes as a companion diagnostic this is a press release from roches ventana in 2013 where they described their intention for developing a companion diagnostic test for insight using iscsi for the small molecule ido1 inhibitor after a couple of years of development work insight decided to compare the specificity and sensitivity of other assays that became available almost five years later in 2018 insight presented this data in asco showing the sensitivity and specificity of rna-scope platform in detection of ido1 expression in tumor biopsies and this was actually after a very short period of development work that we did where we demonstrated that rna scope is about 10 times more sensitive than iscsi that they spent several years to develop and this is an example to re-emphasize that rna scope is much more sensitive than isp and it has significantly faster timelines for development and verification in clinical context i'm going to explain the experimental data in a little bit in in my next slide so basically uh for for that essay halo cells were stimulated with interferon gamma at a varying concentration for 18 hours and asset for idea one expression by either rna scope ish or idea one isp or a western blot analysis the plot on the right here is showing that ifc was 10 times less sensitive as compared to rna scope or our western blot and that it also showed that there was almost perfect correlation between rna scope and western blood sensitivity as you can see the the curve is overlapping and so the strong correlation between rna scope in situ hybridization and western blot so just a highly coordinated induction of ido1 by interferon gamma at both mrna and protein levels finally insight switched from ifcsa to rna scope platform for their clinical trial we performed a reproducibility study for ido1 excuse me shown here is this is the bad graph obtained from idea one rna scope reproducibility resting in melanoma samples and it can be seen here that the results from different instruments as well as those obtained at different time points are highly reproducible and then they used rna scope in a very large clinical study on over 2 500 patients it was used in several large phase 3 studies i'm showing this here to emphasize the fact that rna scopes performed consistently across many large studies involving multiple different tumor types unfortunately the ido1 did not show efficacy in improvement of the patients in terms of response to treatment but the assay itself performed extremely well on a wide range of patients in clinical trials and this idea one study addresses a frequently asked technical question about how rna and protein correlate it showed that rna and protein correlates well when you have equally sensitive assay for detection of rna and protein now switching gear little bit so rna scope is equally relevant in car t therapy and we know that car t therapy has revolutionized the immunotherapy world so the therapeutic car t cells are generally administered systemically and must traffic to metastatic disease site and function within that human micro environment since both are comprised of portions of various human proteins it is almost impossible to distinguish engineered t cells from endogenous cils using antibodies but the arnesco technology utilizes unique reasons for probe design and enables visualization of rt and pcr pcr cell distribution in tumor micro environment while uh corresponding with a marker of activation such as granzyme b or interferon gamma enables assessment of activation states corresponding with a cell type markers such as cd3 or cd4 or any other like cd4 like cd3 cd8 cd4 any any other cell markers it enables assessment of functionality and health monitor anti-tumor activity this is a biodistribution data from ccrp cell therapy in a patient clinical trial conducted by adapt immune nyeso1 is a tumor antigen expressed in many poor prognosis multiple myelomas adapt immune developed a gene modified autonomous tumor reactive pcr against the nyeso1 peptide called sphere they used rna scope to understand mechanism of response and resistance to pcrt cells and they performed ish on ffpe tumor biopsies prior to and following adoptive t-cell transfers in synovial sarcoma and they showed a correlation between the anti-tumor effect and the infiltration of the engineered pcr t-cells into tumors as you can see here the trafficking of tcsc cells is shown in red and seriously shown in in blue this is a data which was presented by bellicam at asco gi recently in 2020 and using rna scope in situ hybridization they observed infiltration of star positive cells into tumor tissues in all all treatment samples whereas endogenous tsp expression and cdc positive lymphocytes remained at the same level in baseline and on treatment samples and bellicon will be using rna scope for their upcoming clinical trial so in short i would like to summarize about affirmative services that we provide scientific pre-sales guidance to help you plan your study with one-on-one study conference to understand your goals our scientists are available to present the summary data via webex or any other video conferencing platform to the key stakeholders our project manager will ensure timely delivery of your study and that objectives are met these are some of the accolades that we have received from our happy clients with this we come to the end of this presentation and uh we will continue to be online for a few more minutes please feel free to send your questions to our panelists who are waiting to answer your queries thank you thank you very much alpana and shauna what an excellent presentation that was from both of you we really appreciate it we do have a couple of questions coming in so um before you leave today please do if you want to please pop the questions in the question box and our team are online actively answering them right now um this has been recorded and so in the next 24 hours this presentation will be available online at our acgbio.com um recorded webinars a page which you can download which you can click and watch again for your enjoyment in case you missed anything do uh do reach out to the pas team and also the support team if you have any questions and if we don't get to answer your questions today we will follow up via email in the next few days with that thank you very much