Deep Insight Into Immuno-Oncology: Immunotherapy Pathways, Targets, and Biomarkers

[Music] welcome to this infographic based educational activity in which dr. Nadim Riyaz from Memorial sloan-kettering Cancer Center discusses the fundamentals of amino oncology including current and emerging cancer immunotherapies and biomarkers to optimize treatment selection for patients with cancer the following podcast is part of a certified educational activity titled deep insight into immuno oncology a visual exploration of current and emerging pathways targets and biomarkers to maximize the potential of cancer immunotherapies access the entire activity and complete the post-test at peer view comm /k and t8 6-0 downloadable infographics and additional resources are also available hello this is Nadine Rios I'm the Associate Director of genomic operations at Memorial sloan-kettering and a physician scientist welcome to module 1 fundamentals of amino oncology so we'll begin with a brief introduction of how the immune system functions in normal physiology as well as how the immune system interacts with cancer the immune system has evolved to help protect our body from pathogens such as viruses and bacteria our immune system is divided into two subsystem the innate and adaptive components these all derive from a common multi in Hema biotic stem cell the innate immune system is the first responder to a wide variety of pathogens as well as danger signals from tissue and include cells such as macrophages neutrophils eosinophils basophils amongst others the adaptive immune system includes things like T lymphocytes and B lymphocytes and generates a response that is more specific to a particular pathogen and can be remembered long term I am memory cells so how do the innate and adaptive immune systems connect together well on the introduction of a pathogen it needs to be detected by some component of the innate immune system often this is by either dendritic cells or other professional antigen presenting cells which might engulf one of these abnormals and components of these pathogens such as in bacteria the cell wall or in viruses certain types of double-stranded DNA will trigger a pattern recognition receptor like a toll-like receptor or C gasping amongst others activation of these receptors will then lead to the generation of cytokines which will recruit the adaptive immune system to investigate what's occurring and subsequently lead to an appropriate adaptive response so how does the adaptive immune system actually generate a more specific response for a pathogen so if we take t-cells as an example each t-cell has a unique t-cell receptor which is responsible for recognizing an antigen that antigen could be a pathogen from a virus or bacteria or that antigen could be something from cancer although we only have 20,000 genes in the genome we have over 10 to the 20th possible t-cell receptors and this diversity is actually generated by a process of vdj recombination so there's a locus in the genome that consists of segments that get rearranged to produce a wide variety of unique t-cell receptors this includes a V segment a diversity segment and adjacent the D segment includes a generation of random sequences which lead to an enormous number of possible recognition motifs for our t-cell repertoire now of course an important role of the immune system besides recognizing an abnormal pathogen or recognizing a cancer is going to be preventing autoimmunity this random generation of t-cell receptors is at risk of leading to autoimmunity and so T cells undergo their primary education in the thymus where they undergo a process of positive selection and negative selection T cells are first selected for t-cell receptors that are able to bind to MHC class 1 to make sure that these T cells have some functionality T cells are not able to do this undergo apoptosis and then T cells had to strongly recognize a self antigen R clonally deleted and also undergo cell death this leaves us with T cells that recognize antigens that are not present in self and can help us identify bacteria other pathogens so how does the immune system play a role in cancer we often thought of cancer as a genetic disease which involved alterations in important cellular processes such as invasion and metastasis evading growth suppressors dis regulating a proliferative signaling and dis regulating cell death however in the past decade or two we've learned through preclinical investigations that tumors also required the ability to evade immune destruction as well as there's a counter side of the immune system which can actually facilitate angka Genesis now we've known for a long time that the immune system is actually prognostic in cancers the increased number of cd8 t-cells and malignancy was prognostic of outcome whether or not patients received immunotherapy so if patients just received surgery or chemotherapy or radiotherapy patients with a higher number of cd8 t-cells had an improved outcome whereas those with higher numbers of macrophages or marlatt drive cells tended to have a worse outcome however it wasn't clear if this was just a creative observation or something that may be causative Lee exploited therapeutically we had a hint though that there was a causative relationship for this observation from evidence from transplant patients patients who undergo organ transplant as everyone is aware receive significant amounts of treatment dampen their adaptive immune system and although initially it was thought that the suppression of the adaptive immune system led to just a virally related cancers more careful epidemiologic studies have demonstrated that dampening the adaptive immune system actually increases the frequency of a wide variety of cancers including lung cancer colorectal cancer skin cancer melanoma amongst others suggesting that t-cells play an important role in immune surveillance and preventing the outbreak of malignancy importantly further refinement the verm histologic understanding of cancers has demonstrated that t-cells are not just randomly distributed in tumors are however characterized in two subtypes some tumors have wide infiltration of t-cells other tumors have t-cells that are excluded from the tumor suggesting the tumors actively evading the immune system then of course some tumors have very sporadic immune infiltrate suggesting the immune system hasn't been fully triggered by the cancer further important role of the attachment immune system and malignancy so with that introduction we'll begin with a brief overview of the innate immune response to tumors these cells play an important role in also facilitating the therapeutic benefit of many of the treatments we use in clinic today so NK cells of course play an important role in f/c mediated antibody dependent cellular cytotoxicity that happens from antibodies that we use such as acetic Simha these tumor infiltrating innate immune cells that are the first responders will also end up secreting cytokines that can alter the chemokine Malou of the tumor and lead to either a tumor promoting micro environment or micro environment they'll facilitate an adaptive immune response going into a little more detail about the natural killer cells these are a group of innate immune cells that show spontaneous cytotoxic activity against cells under stress such as tumor cells and virus infected cells after activation and K cells have an effector immune response by secretion of several cytokines such as interferon-gamma tumor necrosis factor-alpha gm-csf and chemokines i can modulate the function of the innate and adaptive immune cells importantly in K cells play an important role in ensuring that all normal cells are expressing the human leukocyte antigen are able to undergo process of immune surveillance cancer is that down regulate human leukocyte antigen may be susceptible to NK cell based treatments although as we've previously discussed on K cells are considered part of the innate immune system it's clear that some subsets have memory like features that may contribute to an anti tumor response excitingly there are many and k cell based cancer immunotherapies on the horizon including NK cells and cancer immuno surveillance adoptive NK cell therapies cytokine therapies chimeric antigen receptor and gay cell based therapies and then of course monoclonal antibody therapies that target a variety of NK cell checkpoints another important component of the innate immune system is these pattern recognition sensors which first trigger the immune system to know that something abnormal is occurring in the micro environment of a tissue one of the most important of these sensors for cancer is the cytosol DNA sensing pathway called C gas sting historically this pathway was thought detected double-stranded DNA from viruses that might be in the cytosol however it's become clear that other sources of double-stranded DNA can trigger this pathway as well they include things like DNA from mitochondria or DNA caused by genomic instability from cancers and so as we know cancers are quite genomically unstable they often generate double-stranded DNA that leaks from the nucleus to the cytosol they may be activating this pathway activation of this pathway and the generation of interference and an acute setting is thought to facilitate an adaptive immune response however this pathway has another face which is that chronic see gasps ding signaling may actually facilitate the activation of non-canonical NF kappa-b signaling and actually facilitate metastasis and invasion because of these two phases of this pathway this pathway is an active area of investigation for therapeutic manipulation in clinical trials at the moment and this appears to be one of the main pathways by which the innate immune system can identify that there is an abnormal growth occurring although the immune system plays an important role in suppressing cancers if an abnormal inflammatory response occurs it can actually on the other hand that promote malignancy and so we've known for quite a while that tumor promoting inflammation occurs in human tumors during a normal inflammatory response by the unit an adaptive immune system immune cells carry out doesn't need a task of engulfing or destroying foreign invaders however within a complex tumor microenvironment the same infection-fighting immune cells can be reprogrammed by cancer cells and as result instead of destroying the transformed cells these anti-tumor immune cells are subverted into tumor promoting immune cells that can secrete pro survival pro migration and anti detection factors that allow for growth and metastasis how does the adaptive immune system respond to tumors so I think in a very simplified manner t-cells are the main components of the adaptive immune system that can recognize a cancerous foreign and when they recognize a cancerous foreign they can unleash a cytotoxic response I can lead to tumor cell death the best way of understanding this is the cancer immunity cycle initially described by Chen and if we think of tumors in the micro environment cancer cells occasionally undergo spontaneous cell death which leads to the release of tumor antigens picked up by professional antigen presenting cells such as dendritic cells these in verdict cells traffic to the lymph nodes and in the lymph nodes these dendritic cells present antigens to t-cells and lead to t-cell priming and activation activated cytotoxic T cells from the lymph node and travel via blood vessels back to the tumor microenvironment and in the tumor microenvironment these activated t-cells look for the tumor antigens in cancer cells and when they recognize a tumor antigen in a cancer cell they unleash a cytotoxic response on that cancer cell killing that cancer cell so one fundamental step in this cancer musical is a recognition of cancer cells by T cells and this process occurs by an important pathway called the HLA antigen presentation pathway HLA class 1 molecules are presented on all cells of our body these molecules present random proteins on the cell surface and cancers the goal of this pathway is to identify abnormally expressed or perhaps mutated proteins on the cell surface that can be recognized by a t cell as a foreign and so this HLA T cell receptor recognition is considered a signal one to activate a T cell of course T cells need multiple signals for activation so the first step is this MHC complex and then of course T cells also need a CO activation stimulus this is often a cd28 based stimulation and this is often considered signal to T cells that don't have this T cell recognition step ie there is no antigen that the T cell recognizes in the cell is foreign will have no response if there is that first signal but there is no ko activation signal and then T cells will undergo energy now there are multiple classes of antigens that could facilitate the immune system recognizing a cancer as foreign one of the most interesting types of these antigens are neo antigens these are antigens that are derived from mutations as we all know cancer is a genetic disease caused by a number of mutations and these mutations lead to all proteins which can then be presented on the cell surface of tumors to the immune system for the immune system to recognize an abnormality these mutated peptides are not present in normal cells now what's intrigued many with uneo antigens is there's been described a very clear relationship between the number of mutations in a tumor and the objective response rate to checkpoint blockade in clinic and so if these mutated antigens are important one would speculate that tumors with more mutations should be more likely to respond to checkpoint blockade and in fact we do see in clinic that tumors that on average have a higher number of somatic mutations are more likely to respond so T cells which are one of the prime meteors of the adaptive immune response have been an active area of investigation for how to facilitate an immune response in human tumors and their wide variety of activating receptors and inhibitory receptors that can be pharmacologically targeted in a wide variety of these are in clinical investigation today so we can either try and further activate t-cells by facilitating that activation or we can try and remove inhibitory signals from T cells to try and promote activation so how can we target these pathways to trigger cancer immunity well going through the cancer immunity cycle we see there are many points of entrance to try and facilitate an immune response and depending on whether we're thinking about trying to facilitate antigen presentation we can think about mechanisms of trying to trigger a mutagenic CELTA to release antigens we can think about mechanisms a facilitating antigen recognition we can think about mechanisms of trying to facilitate the priming step in the lymph node we can think about mechanisms to facilitate trafficking t-cells to the tumor and then we can think about mechanisms of improving recognition of cancer by t-cells so there are a number of different ways of activating the immune system in which part in the cycle you want to adjust alters how you think about activating the immune system for that specific step I think more broadly we can think of this sort of in three big categories we can think about parts of the immune system where immune system is built our recognize the cancer and get the immune system turned on and that often we can think about things like vaccines or adoptive t-cell therapy or car keys or tills which facilitate turning on the immune system having the immune system recognize that cancer is foreign we can also think about things that turn the gas on on the immune system those include things like cytokines toll receptor agonists and other agonist antibodies and then of course the approach that's had the most success today in clinic is removing the break on the immune systems checkpoint blockade pd-1 and PDL one all right so let's now with that brief understanding of how t cells work in cancer look at how the fundamental checkpoints that are used in clinic today function this animation illustrates how some of the immunotherapy agents called immune checkpoint inhibitors work in the treatment of cancer cancer cells are able to evade recognition by the immune system which allows them to survive cancer cells Express PD l1 which binds to PD one on T cells to turn off anti tumour responses this is called an immune checkpoint immunotherapy prevents cancer cells from inhibiting T cell attack anti PD one antibody therapies blocked the immune checkpoint allowing the immune system to recognize cancer cells anti PDL one antibody therapies can also be used to block this negative interaction immunotherapy allows the immune system to recognize cancer cells leading to anti tumour responses in cancer cell death other immune checkpoint inhibitors act outside the tumor micro-environment in the lymph nodes antigens shed from dying or replicating cancer cells are recognized by antigen presenting cells in the circulation and taken to the central lymphatic system in the lymph node antigen presenting cells interact with T cells via cdat and cd8 e 6 ligands and ctla-4 receptors on t-cells ctla4 is the major negative regulator of T cells which out-compete stimulatory cd28 by binding cdat or cd8 e6 receptors on antigen presenting cells with higher affinity to turn off anti tumour responses this is another immune checkpoint blocking ctla-4 negative regulation with an immune checkpoint inhibitor freeze cdat or cdat six ligands to bind cd28 leading to t cell activation active t cells can then move through the circulation infiltrate the tumor microenvironment and initiate anti tumor responses to kill cancer cells so the primary checkpoint access used in clinic today is the PD one PDL one inhibitory checkpoint this checkpoint plays an important role in peripheral immune tolerance so earlier in this module we discussed central tolerance to try and prevent autoimmunity pd-1 and PDL one play a role in the periphery to try and prevent autoimmunity from occurring when a naive T cell is activated it up regulates PD one and if t cell is able to clear the antigen PT one is down regulated and that T cell turns into a memory cell however if the adaptive immune response is not able to clear the pathogen or the cancer a T cell will become exhausted and upregulate PD one further this PD one PDL one access plays an important role in acute infection cancer and autoimmunity and cancers this pathway is often hijacked by malignant cells to prevent an adaptive immune response from occurring in tumors and so tumor cells often will up regulate PDL one to inhibit T cell activation on a molecular basis we had briefly talked about how the MHC TCR complex or signal one is the first step in T cell activation and that we then needed a CO activation signal or signal - which is based on cd28 binding to one of its ligands such as cdat and downstream activation of the t-cell receptor and this signal to util transcription factors being activated such as n fat now PT 1 and PDL one are actually inhibitory to both TCR and cd28 signaling so PT 1 signaling will dampen downstream effects of TCR signaling as well as dampen downstream effects of cd28 signaling inhibiting t-cell activation decreasing t-cell proliferation and decreasing the likelihood of t-cell survive this pd-1 PDL one interaction can occur either between the T cell and a tumour cell or a T cell and an antigen presenting cell so it's not just limited to the T cell tumor it's all interface although a majority of the activity of pd-1 is thought to occur at the T cell tumor cell interface so immune checkpoint inhibitors primarily functioned by relieving this T cell inhibitory signal the other checkpoint that has been clinically approved is ctla4 and in clinic drugs that target this checkpoint include drugs like aluminum at this checkpoint primarily functions at the T cell engine presenting cell interface and is thought to inhibit T cell priming ctla-4 can bind to cdat or cat6 and function as a negative regulator of T cell priming the use of ctla-4 will inhibit this negative checkpoint and facilitate T cell activation and subsequently facilitate T cell priming and the generation eventually of cytotoxic T cells that can target tumors now importantly this effect occurs in lymph nodes and not in the tumor microenvironment now how about the tumor microenvironment and T regs and how they influence cancer immunity non infiltrated tumors often have poor chemokine expression often have a significant amount of extracellular matrix that inhibits migration of immune cells into the tumor T cell infiltrated tumors on the other hand will often have activation of a wide variety of cytokines that can wind up over the long term resulting in a suppressive micro environment with the expression of a wide variety of chemokines that dampen T cell activation further abnormalities in the micro environment can lead to metabolic dysregulation which can deactivate T cells welcome back to our edge traffic activity and mean oncology will now begin module 2 cancer immunotherapy pin biomarkers the current landscape will begin by looking at immune checkpoint inhibitors immune checkpoint inhibitors have been approved in multiple indications there are three main classes of drugs does that target PD 1 directly that includes nivolumab imp embolism AB those that target PDL 1 that includes a T's ilysm AB vilem a bender rhythm and those that target a different checkpoint access ctla-4 which includes a paluma map and the development of these drugs in the clinical setting is preceded in a typical manner for new drug development in cancer these were first tested in the metastatic and relapse setting and have shown remarkable activity in a wide variety of malignancies and they've subsequently moved into the metastatic first-line setting where their standard of care in that setting again in a wide variety of malignancies and today we're investigating how to use these agents in the most appropriate manner in locally advanced in earlier stage disease will now look at the immune checkpoint inhibitor landscape and specifically selected mono therapy indications pd-1 and PDL one inhibitors have been approved in multiple cancer subtypes these are probably one of the broadest active class of agents since the advent of cytotoxic chemotherapy the other checkpoint actively used in clinic is ctla4 inhibitors which i've been approved in melanoma beyond their broad activity the other exciting thing about anti pd-1 agents in particular is that their responses appeared to be quite durable you see here 5-year overall survival from keynote 0:01 for patients who had either melanoma or non small cell lung cancer results from looking at patients with melanoma we know cell cancer non-small cell lung cancer who received nivolumab show similar results patients with melanoma and kidney cancer having five-year survivals of just under 40% and those would not small cell lung cancer I mean 5-year survival just under 20% and so we see for previously incurable diseases that there are patients with metastatic cancers living out five years suggesting that this response is durable and maybe curative in a subset of patients with their success in the metastatic setting these are now starting to be investigated in the locally advanced setting the Pacific study looked at the use of Davila mAb after standard of care chemoradiotherapy and locally advanced non-small-cell lung cancer you can see the results for progression-free survival demonstrating a significant improvement for patients who received for Villa mAb instead of placebo and again we see almost 10% improvement in overall survival at the two-year time point suggesting a big benefit of Kerala mob in the earlier setting in lung cancer we've seen similar results in melanoma looking at results for checkmate 238 we see in 10% improvement in relapse free survival at two years with the use of Nivola mAb in the adjuvant setting and locally advanced melanomas similar results were obtained in keynote 0-5 for looking at pembo is map in admin settings showing a 20 percent improvement in relapse free survival and so we've seen that these checkpoint inhibitors are not only active in metastatic setting but are now showing signs of activity in locally advanced in earlier disease stage settings I'm moving forward we'll briefly examine a variety of combination strategies for p1 and PDL one inhibitors there are four main categories of combinatorial strategies that have been approved in clinic those include the combination of a PD 1 inhibitor and a ctla-4 inhibitor people have investigated PD 1 and PDL 1 access drugs with chemotherapy and then of course there has been recent interest in looking at combining the PD 1 access with Vega inhibition so looking at IO IO combinations check made 0 6 7 examined the role of nivolumab in a Paloma map combined versus mono therapy with epilim ab or mono therapy with Nivola map numerically this showed that the combination of the follow Bennett and aluminum AB had highest survival according to the updated five-year results from the checkmate 0 6 7 trial both in the volume AB alone or in combination with apollomon map provided significant improvements in overall so in progression-free survival over a plumie map alone in patients with advanced treatment naive melanoma but the combination yielded the most impressive outcomes with 52% of patients still alive at five years the median overall survival was not yet reached for nivolumab plus a paluma map representing the only treatment for metastatic melanoma for which median survival exceeded 60 months and in checkmate 7 we again saw that the combination of Olam haba nipple mmm have improved survival compared to patients who receive chemotherapy the results were from part 1 of the checkmate 2 to 7 trial and wishing the volume AB plus epilim AB met the endpoint of overall survival demonstrating superior benefit versus chemotherapy in patients whose tumors Express PDL 1 greater than or equal to 1% additionally in an exploratory analysis results showed improved overall survival for patients treated with the combination with PDL 1 less than 1% this combination represents a new chemotherapy free first-line option for patients with advanced non-small-cell lung cancer in addition to the approval of new volume AB plus appeal umem AB for first-line treatment of metastatic non-small cell lung cancer expressing PDL 1 greater than or equal to 1% as determined by an fda-approved test and without EGFR or elk genomic tumour aberrations based on checkmate 2 to 7 the combination of new volume AB plus AP Lumia mab into cycles of platinum doublet chemotherapy was also FDA approved this indication is for first-line treatment of metastatic or recurrent non-small-cell lung cancer with no EGFR or ALP genomic tumor aberrations regardless of PDL 1 expression status and is based on results from the checkmate 9la trial the combination of the volume AB a nebula meme AB is fda-approved for intermediate or poor risk previously untreated advanced renal cell carcinoma the approval was based on the results of the check made to one for trial furthermore this ioio combination was recently fda-approved for a Patou cellular carcinoma in patients who have been previously treated with a Serafina approval for this indication was based on the overall response rate and duration of response seen in the combination cohort of the phase 1 - checkmate 0 4 0 trial so moving forward to look at the other two prominent combinations that are being investigated in clinic which include io+ chemotherapy or io+ anti veg f agents so io + chemotherapy has been examined in keynote 189 and keynote 407 both studies looking at metastatic non-small cell lung cancer and both studies demonstrating a significant improvement - amberle is not combined with chemotherapy compared to placebo or a chemotherapy alone and we see in both a significant improvement in overall survival the addition of Pamela's map to chemotherapy impassion 130 investigated the use of the t's ilysm and triple negative breast cancer and again showed an improvement in overall survival in PDL 1 positive patients compared with chemotherapy alone in the metastatic triple negative breast cancer setting anti pd-1 agents have also been combined with veg F directed therapies and so key node 0 46 exam at Pemberley zoom AB with a small molecule t ki exit nib which hits multiple veg F targets and demonstrated that the preamble is imatinib combination had a survival advantage in power 1:50 combined PDL 1 inhibitor with chemotherapy and that Jeff inhibitor and showed that the combination of chemotherapy veg F inhibition and PDL one ambition was superior to veg F inhibition chemotherapy alone suggesting that there may be synergy between remaining PDL 1 based therapies with chemotherapy and an anti veg F drafted treatment by increasing the activity of the immune system immune checkpoint blockade can also cause inflammatory side effects which are often termed immune related adverse effects or Irae s let's look at a brief animation explaining by our AES this animation illustrates the unique spectrum of immune related adverse effects associated with immune checkpoint inhibitors immunotherapy such as checkpoint inhibition enhances the ability of the immune system to recognize cancer cells leading to the anti tumour responses and cancer cell death the enhanced immune activity with immunotherapy is associated with autoimmune responses known as immune related adverse effects immune related adverse effects can affect a range of organ systems including the nervous system cardiovascular pulmonary gastrointestinal musculoskeletal ocular endocrine dermatologic hematologic and renal systems it is important that patients caregivers and healthcare professionals remain vigilant during and after immunotherapy to ensure timely recognition and management of immune related adverse effects so that patients derived maximum benefit from these therapies moving forward what are our current immunotherapy biomarkers so let's start with PDL one expression perhaps the most important biomarker for immunotherapy so importantly PDL one can be expressed on either tumor cells or immune cells PDL one expression is often an adaptive mechanism as cd8 t-cells infiltrate into tumors and cytokines were released this can facilitate the expression of PDL 1 on tumors importantly there are multiple mechanisms of testing for PDL 1 in the tumor microenvironment so each checkpoint inhibitor has its own assay to assess PDL 1 staining here we see a table of the four most commonly used checkpoint inhibitors leading to vomit embolism I have a tease ilysm of interval amad they all have a unique IP based assay with a unique monoclonal antibody that tests for PDL 1 staining in some indications PDL 1 staining is complementary whereas in other indications PDL 1 standing is actually companion diagnostic required for treatment in some indications we only have concerned about PDL 1 thinning on tumor cells specifically and in other indications we care about PDL 1 standing on both tumor cells and and we selves importantly the cut points used are also quite different between the different indications and the different drugs so in summary a PDL one expression is an FDA approved biomarker for member lizhinton non-small cell lung cancer your athili old cancer gastric cancer cervical cancer had a neck cancer and esophageal cancer PDL one singing is also approved for guiding the t's loosen up treatment in triple negative breast cancers and yura thelia malignancies based on the most recent FDA approvals PDL 1 expression testing is also required to guide treatment decisions for the use of a t's ilysm ab mono therapy as well as the combination of new volume AB in appeal umem AB in the first line setting of non-small cell lung cancer here we see data for outcomes in keynote 0 for 2 by PDL one level this is a population of non small cell lung cancers comparing patients who receive Pember Elizabeth versus chemotherapy and we can see in patients with PDL one sitting over 50% there's a significant improvement in overall survival for patients received Pember lisanna with patients with PDL one scene between 1 and 49% there appears to be no difference between the Pemble asthma and chemotherapy arms and overall survival we see very similar results in keynote 189 which looked at member lizmac combined with chemotherapy versus chemotherapy alone and we see again that is the fraction of tumor cells that have higher PDL one seeing increases the benefit of adding embolism at further increases similarly in impassion 130 which looked at the use of at easel is a naba to chemotherapy and with metastatic triple negative breast cancer we see that in the PDL one positive patient population there's an improvement in overall survival which does not exist in the overall population I'm moving on to the next fda-approved biomarker which is microsatellite instability so microsatellite instability is a deficiency of the MMR DNA repair pathway there's a hereditary syndrome called Lynch syndrome which results in hereditary defect in one of these genes which predisposes patients to rectal cancer endometrial cancer can also occur sporadically due to sounding of mlh1 due to promoter methylation there are multiple strategies to test for this routinely in clinic including immunohistochemistry for components of the mismatch repair complex including mlh1 pms - MS h2 and ms h sex or microsatellite instability testing by pcr looking at five different low sigh you can also use next-generation sequencing to identify tumors that are microsatellite unstable Luis Diaz and other investigators showed that patients with mismatched repair deficient colorectal cancers had improved overall survival after receiving anti pd-1 therapies and they subsequently demonstrated that this was a pan-cancer phenotype so regardless of histology tumors that were mismatched repair deficient had a significantly high response rate to anti pd-1 treatments so microsoft instability is an fda-approved biomarker for parallelism AB in solid tumors of any histology and colorectal cancer and for the use of new Volm AB a new volume AB plus a Paloma mAb in metastatic colorectal cancers that are microsatellite unstable moving forward with our next current immunotherapy biomarker tumor mutational burden so tumor mutational burden as we've discussed in the prior module is considered a biomarker for immunotherapy because it may indicate how foreign a cancer appears to the immune system so these mutations that are generated and often they're passenger mutations result in changes in proteins which are subsequently expressed or presented on the cell surface by MHC class 1 and that can be recognized by a t-cell as foreign initial data demonstrating that tumor mutational burden was associated with outcomes originated from Memorial sloan-kettering looking in the patients with melanoma received anti ctla-4 therapy and this study demonstrated that patients who had a long term benefit to anti ctla-4 therapy had a higher overall number of coding mutations and that the survival of patients with over 100 mutations of their exome was markedly improved compared to patients who had less than 100 mutations in our Excel this was similarly reproduced in a cohort of patients with small cell lung cancer again demonstrating improved rates of patients with durable clinical benefit who had higher TMP compared to those with lower can be interestingly many paces with higher TMP had mutations in DNA repair genes and importantly as we've seen previously the response rates in Phase one to two studies across cancer types to anti pd-1 therapies appears to be directly proportional to the median number of somatic coding mutations in a tumor type so cutaneous squamous whole cancers Geoff one of the highest tumor mutational burden is due to their marked exposure to UV light have one of the highest response rates to anti pd-1 therapies whereas pancreatic cancers which have a relatively lower tmb have a lower rate of response in the phase one studies tumor mutational burden is a continuous variable and obtaining a cut-off can be challenging a wide variety of cut-offs have been proposed data from checkmate 2 to 7 suggested tumor mutational burden of around 10 may be optimal in check made to to 7 patients with a tumor mutational burden greater than 10 appeared to have a higher progression free survival and they received nuvola mandible mmm compared to chemotherapy however the results of the overall survival analysis did not suggest that this can be cut off was necessarily predictive for benefit mystic was a study after vilem AB + tremolo versus chemotherapy versus der villain abalone in metastatic non-small cell lung cancer and in this study the authors investigated the use of a blood based assessment of tumor mutational burden and its association with outcomes they used a slightly higher cut point than what was previously discussed cut point of 16 and they identified that a blood based TM be greater than 16 was associated with benefit for gervilen have intramural mmm compared to chemotherapy in the high tmb group whereas in the low tmb group outcomes were roughly similar between the three treatment arms again PDL one expression in tumor mutational burden appeared to be quite independent of each other this is data from sloan-kettering demonstrating the lack of any correlation between the two suggesting they provided independ information how can we measure tumour mutational burden in the clinic many of the studies we've just discussed are research studies based on whole exome sequencing which sequences 30 mega bases of the coding genome there are of course panel based mechanisms that are often used at and if I targeted therapies that can also give us information about too many mutational burden those include things like a panel from Foundation at Memorial sloan-kettering we have our own in-house panel evaluates coding genome called msk CCD impact although all of these methods can identify tumor mutation burden we need a conversion factor between these different methods because the depth of sequencing is different between them the amount of the genome the sequence is different between them and so their precise number that they're going to provide for the tumor mutation burn estimate it's going to be different between different panels and so if there's ongoing effort by a friend's of cancer research to try and harmonize tumor mutational burden between the different panels once these tumor mutational burden numbers harmonize between different panels and we need to think about what is a good cut point to distinguish high and low tumors and as we had previously discussed identifying high and low tumors can be challenging as tmb is a continuous variable welcome back to module three of our educational infographic activity this module will be on cancer immunotherapy and biomarkers the next wave of progress well first begin with new and emerging immunotherapies in oncology so there are a wide variety of combination based studies ongoing in clinic today over a thousand studies investigating combination of anti pd-1 with a broad spectrum of agents the most popular classes of combination strategies we previously reviewed including the combination with chemotherapy with other IO agents such as ctla-4 and with drugs that target the veg F access however there are innumerable other targets being looked at both as monotherapy and in combination with anti PD one therapy here we'll review some of the most exciting possibilities for future treatments well first look at the agents that target the micro environment and regulatory mechanisms of the immune system as we briefly discussed previously and the veg F access appears to be synergistically functional with the immune system that Jeff signaling is often aberrantly regulated in tumors and can lead to abnormal vasculature as well as stimulating myeloid Drive suppressor cells and type hf therapies can subsequently normalize the vasculature and lead to enabling a recruitment of t-cells into the tumor microenvironment and so this is currently an active area of investigation some combinatorial approaches with anti-angiogenic therapies have already translated to clinical success and regulatory approvals but many other studies examining the volume AB at Ezell ISM AB a volume AB the Lummi map and other checkpoint inhibitors paired with anti veg F monoclonal antibodies or anti-angiogenic T key eyes are underway IDO is an enzyme that metabolizes tryptophan and is present in tumor cells dendritic cells macrophages and B lymphocytes and in preclinical models inhibition of IDO leads to facilitation of an immune response idea itself supports an immunosuppressive tumor micro-environment there was widespread enthusiasm based on preclinical data that this may be applicable in humans unfortunately the clinical results are not as promising the investigation of epic a tostada an idea 1 inhibitor tip embolism AB demonstrated no improvement in progression-free survival and so this is sort of a cautionary tale of the difficulties we have in translating findings from preclinical systems and murine models of cancer immunology to humans as in there are significant differences between the murine immune system and the human immune system will move forward to sell our based therapies these include both adoptive T cell based therapies and engineered T cell based products adoptive T cell therapies have been pioneered by people like Steve Rosenberg at the NCI and involve extracting T cells from individual patients tumors expanding them ex vivo to those T cells that react best with tumors and then re-injecting them into patients this has had promising activity in limited numbers of patients and a constellation underway to try and further improve on these initial results in addition to using a patient's own T cells expanded we can engineer T cells to specifically react to tumors I perhaps the most exciting example this has been the generation of car T cell therapies and liquid malignancies and the way these treatments work is patients with relapsed refractory b-cell malignancies have T cells extracted from them by Luca phoresis those T cells are then transferred by retro viral or lentiviral transduction with anti cd19 engineered t cell receptor this T cell receptor includes an antibody fragment that recognizes C T 19 engineered on to a t-cell receptor that's activated when this binds to CD 19 these genetically engineered T cells are subsequently isolated and they re infused into a patient after our preconditioning regimen multiple anti cd19 car T cells have been approved for use in human patients including the first in a week lapsed or refractory al L in pediatric or young adult patients subsequently multiple anti cd19 car T cells have been approved in both adult and pediatric B cell based malignancies Carr t-cells targeting other b-cell base antigens such as PC ma are in development and appear promising for anti tumour based activity the next class of therapies will talk about our intra tumor therapies and agonists of the immune system including on coletek viruses and stimulating cytokines uncle it advises our viruses that are engineered to selectively replicate in tumor tissue and subsequently lead to tumor cell lysis that tumor cell lysis is hoped to facilitate the generation of a systemic tumor immune response that can then subsequently lead to the eradication of distant cancer cells there a wide variety of oncolytic viruses being investigated in clinic using a wide variety of ours families Tyvek which is perhaps one of the farthest along in clinical development uses a herpes virus but there are other agents being investigated today including using other viral families T Beck demonstrated in phase 3 trial the optim study improvement in overall survival over gm-csf studies are ongoing looking at t vac in addition to anti p1 therapy and this may be a promising approach of trying to facilitate a recognition step for tumors in combination with anti pd-1 based treatments indeed t vac in combination with epilim ab has demonstrated greater efficacy than either t vac alone or epilim Emad mono therapy and a study has completed accrual looking at t vac in combination with PD one pathway blockade il-2 is well known cytokine that stimulates T cell proliferation induces generation of cytotoxic T lymphocytes and facilitates maintenance of natural killer cells I'll too is also involved in the maintenance of T regs an elimination of self reactive T cells nectar 214 is a CD 122 biased agonist that's pegylated pegylated formulation allows for a slow release of interleukin 2 after IV injection and biases binding of the drug to the interleukin 2 receptors that are more commonly expressed on cytotoxic lymphocytes the biased agonist is less likely to bind aisle to receptor alpha which is responsible for activating t resins this agent is thought to perhaps be more specific for activating cd8 t-cells and activating T regulatory cells phase one data showed promising activity in combination with anti pd-1 therapies and we're now waiting Phase three results in multiple malignancies for whether or not this agent adds benefit on top of anti pd-1 therapies il-2 is indeed an important target and extensive clinical investigations with NK t are to14 also known as Ben Pegg Aldous Lucan orb MPEG + Novoa map are underway these include registration 'el trials in first-line metastatic melanoma cisplatin ineligible metastatic euro Thiele cancer and metastatic renal cell carcinoma as well as an adjuvant melanoma andddd muscle invasive bladder cancer vaccine are another exciting area of investigation and these include vaccines that target shared antigens or vaccines that can target neo antigens so the idea behind a vaccine is that a vaccine is delivered to a patient against a specific antigen that's then taken up by dendritic cells in the patient and that stimulates the generation of a t-cell response that's specific for that patient's tumor and then can lead to tumor cell eradication one of the most exciting recent developments is the development of personalized neo antigen vaccines this involves tumor procurement so taking its sample of the patient's tumor and having that tumor undergo a whole exome sequencing and RNA sequencing to identify neo antigens or mutation based antigens that are expressed in the tumor which is determined by RNA sequencing and that can also bind to the patient's hla class 1 molecules which is determined by HLA typing once mutations that both bind to HLA plus 1 and are expressed or identified a vaccine that specific to a patient can be generated there multiple methods of generating this vaccine whether it's through the generation of synthetic long peptides or mRNAs and there are multiple advents that can be administered with this vaccine including poly IC LC 2 preliminary studies demonstrated that these vaccines can successfully generate antigen-specific responses a study by us on used synthetic RNA vaccines encoding 527 more neo antigens and demonstrated responses in handful of patients similar work was done at dana-farber using along synthetic peptides which suggest a delay of recurrence in a small number of patients studies looking at both synthetic long peptides and RNA vaccines are ongoing to see if these add benefit in addition to anti pd-1 therapies how about combining a type ii d1 with other checkpoints or activating the innate immune system so we briefly review that there are other t-cell checkpoints that can be activated many of these been investigating preclinical systems including molecules that can activate T cells including ox40 Gitter amongst others and then of course other inhibitory receptors like Tim 3 Vista and like 3 it appears that there dysregulation in cancer is more complicated than what the preclinical model suggested and so this is still gonna require some more scientific work before these will be ready for clinical investigation for example real island map a monoclonal antibody that targets like 3 has been studied in combination with nivolumab in patients with melanoma who progressed during prior anti pd-1 PDL one therapy and has shown some activity but further biomarker investigations are needed to determine the best way to take this and other new checkpoint based approaches forward this is important because research suggests that there could be a relationship between expression of targets such as lag 3 as well as their inhibition and likelihood of response in the reliable map studies the response rate was higher in patients with baseline expression of lag 3 research is ongoing to further optimize current checkpoint inhibition approaches including ctla-4 blockade one such approach is focused on regulating the degree of immune activity using nan few isolated antibodies to deplete T regs another approach uses Pro antibodies that can improve ctla-4 blockade specificity by reducing antibody binding outside the tumor micro-environment which helps spare healthy tissues there are a wide variety of drugs turning the innate immune system including as we discussed in module one looking at sting and see gas which are members of the innate immune system that wind up triggering initial interferon signalling of a tumor there put agonist and antagonist looking at that pathway there are also checkpoint inhibitors targeting specifically innate immune system including inhibitory checkpoints on K cells on macrophages and on other members of the innate immune system that are in clinical investigation for example t-cell immune or receptor with immunoglobulin in ITIM domains digit is an immune checkpoint receptor on cytotoxic memory and regulatory T cells as well as natural killer cells which can suppress T cell activation and promote t-cell exhaustion inhibition of digital own or in combination with other checkpoint inhibitors may increase cytotoxic T cell proliferation and function as another example chemokine receptors 2 and 5 on T cells regulatory T cells monocytes myeloid derived suppressor cells and tumor associated macrophages can promote trafficking of the immunosuppressive cells and suppress t-cell function inhibition of CCR 2 and 5 may decrease tumor infiltration of immunosuppressive cells and is under investigation preclinical and clinical studies support ongoing investigations of CCR 2 and 5 dual antagonism so moving forward let's discuss novel biomarkers for amino therapies and so there are three classes of biomarkers those that reflect what's happening in the micro environment those that reflect that what's happening the tumor and those that reflect parts of the host future work will involve refining are currently available biomarkers we previously discussed improving tumor mutation brain by identifying the appropriate cut point and perhaps considering obtaining it from circulating tumor DNA and future biomarkers we'll also consider how to integrate our known biomarkers together including PDL one saying can be an inflammation but of course there are also going to be novel biomarkers including immune cell markers and additional patient features that remain to be uncovered the past few years have seen an explosion of the identification of immune some marketers with single-cell sequencing technologies which will briefly review shortly first we'll discuss emerging genetic biomarkers of response to checkpoint blockade perhaps the most interesting of these is the human leukocyte antigen HLA class one as you recall is expressed on all cells in our body and is responsible for presenting antigens to cd8 cells there are three main classes of HLA class one genes AP and C these genes are the most polymorphic genes in the human genome meaning these genes are the most unique in the human genome we inherit two copies of each of these genes one from mom and one from dad and so we have six unique copies of HLA class one genes for some of us mom and dad are a little bit more related than they should be and so instead of inheriting six unique copies we might inherit only five or four unique copies of HLA class one and so if that happens the number of peptides we can present on our cell surface is going to decrease because the diversity of our HLA class 1 molecules is decreased and so you might anticipate that patients who have less HLA class 1 diversity you're gonna have in fear response to checkpoint blockade that are being in fact that is what's seen both in published exome data and in our own institutional data at sloan-kettering similarly we know tumors are quite copy number unstable HLA a B and C are all located on chromosome 6 and sometimes tumors will lose a copy of either mom or dad's chromosome 6 and when this happens we have to see a very similar effect that patients have an inferior outcome they have only three unique HLA class 1 molecules so we've learned in the past few years that oncogenes and tumor suppressor genes in addition to their role on solid hominis growth can influence the immune micro environment of tumors perhaps the best characterized STK 11 or lkb1 which is known to lead to an immune desert phenotype and not small cell lung cancer similarly Mik and beta contingent or both also known to have immunosuppressive effects on the tumor microenvironment so what about biomarkers in the micro environment such as biomarkers that indicate inflammation perhaps one of the best described inflammatory signatures is an interferon gamma signature which examines the expression of o Friday of cytokines and other molecules associated with activation of interferon gamma in the micro environment this interferon gamma signature can be measured by either RNA sequencing or by using a nano string based platform and this interferon gamma signature has been previously associated with a response to anti pd-1 therapies in phase one studies an 18 gene based gene expression program looking at interferon gamma based genes showed an AUC hit 0.75 for responders versus non responding patients however beyond analyzing at the expression state of a bulk tumor we can use single cell sequencing technologies to look at both the expression of individual cells as well as the antigen specificity of an individual cell so as you'll recall t-cells have an antigen specificity to turn by the t-cell receptor which undergoes vdj recombination and this provides a barcode on an individual T cell T cells also have functional States and generally are thought to include naive T cells which have not been exposed to an antigen which can either mature into effector T cells and a memory T cell population or if they're chronically exposed to antigen can differentiate into exhausted T cells now this T cell state can be determined by the RNA in an individual T cell so by using single cell RNA sequencing we can determine a barcode of a T cell which determines its specificity as well as identify what state that T cell is hen so one of the most interesting recent studies was from Don Schumacher which looked at excising a tumor and investigating whether T cells within this tumor were all reactive to the tumor itself so they isolated cd8 T cells extracted that T cell receptor and then transfected these into T cells to identify how many of these T cells in the tumor actually responded to the tumor and to the surprise only one of the T cells they extract for the tumor was actually reactive to the tumor itself in fact three of the T cells they extracted from the tumor reactive to EBV as measured by interferon gamma on stimulation of struck from the tumour suggesting that most of the T cells in the tumor are actually passengers and not able to recognize the tumor itself this work was partly validated by work from Stanford which used single cell sequencing to look at T cells from patients who received checkpoint blockade both pretreatment and post treatment single-cell RNA sequencing was used to identify different T cell populations and what was interesting was that T cells that were expanded in tumors post treatment actually were not present in tumors pretreatment when we matched samples based on that T cell barcode suggesting that many of the T cells that mediate response for checkpoint blockade are actually not pre-existing in the tumor micro-environment but are recruited from the periphery we ourselves have shown that T cell response in individual patients is directly proportional to the number of neo antigens lost so the number of unique T cells that expand on therapy in patients who respond is linearly proportional to the number of new antigens that become undetectable in therapy suggesting that the T cells are responding to the underlying mutations in a patient's tumor although we've long known that cd8 T cells associated with a response single cell technologies have allowed us to identify subsequent features of these cd8 t-cells that mediate response work near our cones group demonstrated that there were two subpopulations of Seanie eight cells present and individual tumor what they labeled cd8 G appeared to be higher and responder stand-in on responding melanoma patients who received checkpoint blockade therapy and CAG signature appeared to be primarily driven by a transcription factor called TC f7 which mediates a stem like cd8 t-cell state or as naive cd8 t-cell like state and this was validated by IHC and immunofluorescent techniques in a separate cohort sustaining with a both T E and T cf7 identified CET cf7 positive t-cells were higher in responders versus non responders and then subsequent melanoma cohort which has now been validated by several other groups as well suggesting that TC of seven piece ta t-cells may be more important for mediating response than just the overall cd8 t-cell population and other features of the microenvironment may indicate an active immune response so although there's been significant focus on t-cells some tumors have the generation of tertiary lymphoid structures or lymphoid like structures in the microenvironment and so here is a melanoma patient at sox10 highlighting the melanoma cells and here you can see cd3 highlighting t-cell it cd20 highlighting b-cells suggesting the formation of a tertiary lymphoid structure tertiary lymphoid structures and the presence of b-cells has been associated with a superior prognosis regardless of treatment and so these melanoma patients were treated prior to the advent of checkpoint blockade we can see those that have high cd8 cells plus these b-cells suggesting we have the formation of these Trichet lymphoid structures have the most improved outcomes and then lastly there are host related factors that can be associated with outcomes Tim you know therapy and perhaps most interesting of these include the microbiome the microbiome can be obtained from us tool sample on 16s ribosomal RNA sequencing to identify the populations of bacteria that are present in individual patient and microbiome studies have identified taxonomies that associate with patients who respond to treatment compared to patients who don't respond to treatment to date however some difficulties with the microbiome studies have been that the microbiome is quite dependent on the region of the world you live in in terms of which organisms you're exposed to and so comparing studies from different parts of the world have been challenging as unique bacteria have been identified in different studies as being associated with response however important mechanistically fecal microbiome transplant in murine models has been significantly associated with proof response anti p1 therapy so in summary there are many emerging treatments in immunotherapy there are a wide variety of combinations being explored in clinic today in a wide variety of new agents being investigated including agents at target innate immune system like suggesting agonist agents at target other immune checkpoints new engine vaccines and engineered T cells on the biomarker side of things new interesting avenues including many fundamental insights flowing from single cell sequencing of the Myanmar environment as well as netic and host related biomarkers in under active investigation thank you for listening download materials and complete the post-test for instant credit at pier view comm /k n t eight six zero this activity is supported through an educational grant from bristol-myers Squibb this activity has been jointly provided by Medical Learning Institute incorporated and P VI P review 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Length: 63min 39sec (3819 seconds)

Published: Tue Jun 02 2020