>> GOOD AFTERNOON AND WELCOME TO THE PROTEOMICS INTEREST GROUP LECTURE SERIES, THE FIRST IN THIS NEW YEAR 2019. OUR SPEAKER TODAY IS DR. NATHALIE AGAR FROM HARVARD. SHE DID HER Ph.D. IN CANADA AT CONCORDIA UNIVERSITY AND DID A POST DOC AT MARGILL AND HARVARD IN NEUROSURGERY AND THEN GOT THE POSITION IN HER OWN LAB AT HARVARD IN 2002 AND NOW, SHE HAS AN IMPRESSIVE RANGE OF TITLES IN THE DEPARTMENT OF NEUROSURGERY AND RADIOLOGY SHE'S ASSOCIATE PROFESSOR AND SHE'S WORKING ON REALLY CUTTING EDGE APPLICATIONS OF MASS SPEC IMAGING TO THE CLINIC TO ACTUAL NEUROSURGERY IN THE OPERATION, IN THE SURGERY ROOM, SO IT'S REALLY EXCITING AND I'M SURE YOU WILL ENJOY HER TALK. >> THANK YOU VERY MUCH ALEXANDRA AND THANK YOU FOR THE INVITATION AND THE HONOR TO COME PRESENT HERE. SO BEING A PROTEOMICS GROUP, WHILE I DO HAVE TO FIRST MAKE ALWAYS THIS DISCLOSURE PER OUR INSTITUTION BUT THE OTHER DISCLOSURE IS THAT THERE' NOT MUCH PROTEIN WORK, I'M SORRY ABOUT THAT. I MADE SURE TO INCLUDE ONE APPLICATION THAT HAS PROTEIN ANALYSIS AND I GUESS GIVEN THAT THERE'S NOT TOO MANY OF US, I WOULD LOVE IF WE MAKE THIS MORE OF A DISCUSSION, SO IF YOU'RE--IF THERE ARE QUESTIONS ALONG THE WAY JUST CUT ME OFF AND I'M HAPPY TO TAKE QUESTIONS AND WE--I WON'T GO OVER, WE CAN STOP IT WHEN THE TIME IS UP. SO THIS AN OPERATING ROOM IN THE BUILDING WHERE MY LAB USED TO BE UNTIL ABOUT A COUPLE YEARS AGO AND THIS IS IN 1923 AND IT'S A PRETTY HUMBLE ROOM AND IT'S QUITE MIND BOGGLING TO THINK THAT WE WERE ABLE TO DO SURGERY WITH SUCH LOW TECHNOLOGY AND THIS IS THE OPERATING ROOM THAT WE'RE CURRENTLY WORKING IN AT THE BRIGHAM WOMEN'S HOSPITAL, AND WE CALLED MULTIADVANTAGE GUIDED OPERATING SUITE. SO THIS ROOM--I GUESS THE LASER WILL WORK BETTER THAN FINDING MY CURSOR SO THIS OPERATING ROOM IS A THREE ROOM--IT'S A THREE ROOM OPERATING ROOM ON ONE END WE HAVE A CEILING MOUNTED THREE TESLA MRI THAT COMES TO THE PATIENT HERE WHEN NEEDED AND WE CAN ALSO BRING THE PATIENT INTO A PET CT SCANNER HERE AND WE HAVE A RANGE OF DIFFERENT IMAGING TOOLS AND THE MAIN DIFFERENCE BETWEEN THIS OPERATING ROOM AND THE ONE I'VE SHOWN YOU IN 1923 IS THE AMOUNT OF TECHNOLOGY THAT WE'RE BRINGING TO BE ABLE TO VISUALIZE THE SURGICAL FIELD AND TO UNDERSTAND WHAT IT IS THAT WE'RE DEALING WITH DURING SURGERY AND IN THE CASE OF CANCER, IS THIS CANCER OR NOT CANCER? AND SHOULD IT BE REMOVED? SO DESPITE ALL OF THIS TECHNOLOGY, THERE'S STILL--THERE WAS STILL NOT A TOOL THAT BASICALLY ASSESSES THE TISSUE ITSELF AS YOU'RE GETTING CLOSER OR TOWRCHING IT AND THIS IS WHERE WE BROUGHT IN THE MASS SPECTROMETER, SO IN THIS CASE, WE DO FAIRLY SIMPLE MASS SPECTROMETRY, WE HAVE AN ION TRAPPED INSTRUMENT AND THEN IT'S WITH AN IONIZATION SOURCE AND IT MAKES THE GREAT ADVANTAGE OF IONIZATION HAS WE REQUIRE VERY LITTLE TO PRACTICALLY NO SAMPLE PREPARATION. BUT WHAT WE ALSO DID WAS WE BROUGHT THE CHEMIST TO THE OPERATING ROOM, THE PATHOLOGIST, AND EVEN A COMPUTER SCIENTIST HERE IN ADDITION TO THE SURGEON TO BE ABLE TO DEVELOP THOSE APPLICATIONS. SO THE RANGE OF APPLICATION THAT WE'RE DEVELOPING FOR SURGICAL PATHOLOGY AND ONCOLOGY SPANS FROM SECONDS TO MINUTES AND HOURS. THE FIRST ONE IS TO BE ABLE TO LOOK AT SURGICAL MARGIN AND TO PROVIDE THIS FOR THE SURGEON TO MAKE THE DECISIONS. THE OTHER TYPE OF INFORMATION IS TO BE ABLE TO PROVIDE KIAG NOOF THETIC OR PROGNOSTIC INFORMATION SUCH AS TUMOR TYPE, GRADE, BEHAVIORS SUCH AS RECURRENCE OR PROGRESSION AND EVENTUALLY WHAT WE WOULD LIKE TO BE ABLE TO DO IS PREDICT RESPONSE TO TARGETED THERAPY. SO HERE--THERE IS SOME OF THAT INFORMATION THAT WE CAN PROVIDE WITHIN SECONDS BUT WE CAN ALSO GO ALL THE WAY TO A FEW MINUTES HERE AND THE OTHER LINE OF WORK THAT WE DEVELOPED WHICH I USED TO THINK WAS VERY SEPARATE FROM ALL OF THAT DIAGNOSTIC WORK IS TO LOOK AT DRUG DISTRIBUTION AND RESPONSE AND WE STARTED OFF IN PRECLINICAL ANIMAL MODELS WITH AN EMPHASIS ON BRAIN PENETRANTS AND WE'VE SINCE MADE THOSE APPLICATIONS QUANTITATIVE AND ARE APPLYING THIS IN CLINICAL TRIAL ASKS USING THE WORK FLOWS WE DEVELOPED HERE TO BE ABLE TO LOOK AT DRUG DISTRIBUTION IN THE CONTEXT OF THE 3D ANATOMY OF THE TUMOR AND THE PATIENT'S BRAIN. SO THE CLINICAL TRANSLATION OF THOSE TOOLS, HOWEVER, THE TIME SCALE IS IN YEARS, SO THE MOST MATURE PROTOCOL THAT WE HAVE IS FOR BRAIN WHERE WE HAVE AN IRB APPROVED PROTOCOL TO SUPPORT GLEEL CELLSOMA, SO IN THIS CASE, WE PROVIDE THE INFORMATION TO THE SURGEON IN REALTIME AND SHE CAN DECIDE WHAT TO DO WITH THE INFORMATION AND WE'RE EVALUATING HOW TO BEST COMMUNICATE THE INFORMATION AND WHAT--AND SHE'S EVALUATING HOW TO USE THE INFORMATION. THE OTHER LINE OF APPLICATION IS IN BREAST CANCER WHERE WE HAVE--AND I HAVE TO SAY THAT PROBABLY TO CUTS CLOSE TO TEN YEARS TO COME TO SOMETHING LIKE THIS, NOT QUITE TEN YEARS, NOW IN BREAST CANCER WE'VE BEEN DABBLING WITH THIS FOR MAYBE SIX YEARS, NOW WE'RE MIDCLINICAL TRIAL WHERE WE CAN USE WHILE WE'RE EVALENTINEDIAT UG THE POTENTIAL OF USING MASS SPECTROMETRY--PLESES YOU--TO EVALUATE SURGICAL MARGINS IN SURGERY AROUND THE LUMP ECTOMYOSIN ME AND NOW THE RECENT WORK, OUR FIRST REAL PAPER WAS ACCEPTED TODAY JUST BEFORE I CAME HERE IS ON PROSTATE CANCER WHERE WE DO PROOF OF CONCEPT STUDY FOR TUMOR DETECTION BUT ALSO IT SEEMS LIKE WE WOULD BE ABLE TO EVALUATE THE DPLEESON SCORE AND THAT WOULD BE ABLE TO GUIDE MRI BIOPSY AND IN THIS THE MORE PRELIMINARY WORK WHERE WE HAVE JUST VERY PRELIMINARY DATA BUT VERY SIGHTING IS IN LONG CANCER TO BE ABLE TO SUPPORT SURGICAL DECISION MAKING. SO EVERYTHING ELSE IS ABOUT THE BRAIN WHICH IS WHERE WE ARE DOING MOST OF OUR WORK, THE WORK WE DEVELOPED TO DATE USING MASS SPECTROMETRY AS A TOOL IS NAVIGATION WHICH IS SORT OF A GPS FOR THE BRAIN AND AT THE BEGINNING OF THE SURGERY WE REGISTER FEATURES FROM THE PATIENT'S HEAD TO THE PREOPERATIVE IMAGING AND BEING MR BASED OR PET OR INVIVO IMAGING, EVERY POSITION THAT WE TAKE A SPECIMEN AND I SAY WE, I DON'T PERFORM SURGERY. IT'S A TEAM EFFORT. EVERY POSITION THAT WE TAKE A SPECIMEN, WE REGISTER WHERE WE TOOK THE SPECIMEN IN THE 3D ANATOMY AND EACH SPECIMEN WE REQUIRE A FULL RANGE OF MASS SPEC INTEREST BUT WE ALSO TAKE A PESMEN TO THE LANLAB WHEN WE'RE DEVELOPING APPLICATIONS WHERE THE PATHOLOGIST CAN EVALUATE THE HISTOPATHOLOGIES TO BE ABLE TO VALIDATE THIS. SO WE'RE WORKING TOWARDS CONVERTING EXISTING SURGICAL DEVICES SO WE CAN SEND SPECIMENS DIRECTLY TO THE INSTRUMENT BUT THE CRITICAL COMPONENT IS TO BE ABLE TO MAKE THAT WORALATION HERE, IF YOU USE MASS SPECROMETERS YOU WILL ALWAYS GET SOME KIND OF A SIGNAL BUT WHAT DOES IT REALLY MEAN. SO THE TYPE OF IONIZATIONS WE'RE USING WE STARTED OFF WITH DEZI, WHICH STANDS FOR DESSORRAL SPECIFICATION, YOU HAVE A CHARGE SPRAY OF SOLVE EPT THAT HITS THE SURFACE AND THE MOLECULES COME OFF, ABSORB AND ENTER THE INSTRUMENT.& WE CAN ALSO DO IMAGING WHERE WE ROGSTER OR SPRAY IT OVER THE TISSUES SO WE CAN ACQUIRE A TWO D IMAGE TO GAIN HISTOPATHOLOGY. SO WE STARTED OFF WITH BE SI, BUT WORKED WITH THE BRAIN BUT DIDN'T WORK WHEN WE WENT TO BREAST CANCER. THE BREAST CANCER TISSUE IS EXTREMELY FATTY AND AS SOON AS WE PUT IT ON THE SURFACE IT MELTS AWAY AND THE SPRAY, THE ANGLE OF THE SPRAY OF THE SOLVENT WAS JUST PUSHING THE TISSUE IS WITH ABSORPTION SURFACE LIQUID EXTRACTION AND IN THIS CASE, THIS IS THE ONE WE HAVE WHERE WE HAVE A DUAL DESI OR THE FLOW PROBE HERE, YOU HAVE A PROBE THAT COMES TO THE SURFACE OF THE TISSUE, DELIVERS TO SOLVENTS AND YOU HAVE A CONTINUOUS EXTRACTION WHERE THE ANALYTES COME INTO THE MASS SPECTROMETER SO THIS IS WHAT WE'RE USING CURRENTLY AND THE OTHER VERY PROMISING PLATFORM THAT WE WOULD LIKE TO BE ABLE TO BRING AFTER A LITTLE BIT OF ENGINEERING IS LIQUID EXTRACTION SURFACE ANALYSIS AND IN THIS CASE, YOU HAVE A LICK EDUCATIONAL MICROJUNCTION FORMED HERE BUT THIS IS A ONE DROP AND WE CAN GET PRETTY CONCENTRATED AMOUNT OF THE ANALYTE HERE DEPENDING ON THE TYPE THAT WE LEAVE THE DROPLET ON TO THE SURFACE. SO THIS IS BASICALLY THE VERY INITIAL DATA THAT WE HAVE WHERE WE TOOK SOMETHING LIKE TEN SPECIMENS AND ACQUIRED THE SPECTRA FROM EACH OF THOSE SPECIMEN, DID A HISTOPATHOLOGY, EVALUATED THE TUMOR CELL, CONCENTRATION AND TO ME IT ALL MADE A LOT OF SENSE, I WAS VERY EXCITED BUT NOT SO MUCH MY COLLEAGUES AND I WILL SHOW YOU HOW WE CAN VISUALIZE THAT DATA BUT THERE ARE TWO DIFFERENT WAYS WE CAN WORK WITH THE DATA, ONE IS WE CAN USE THE FULL SPECTRUM AND EXTRACT SOME MOLECULAR FINGERPRINT USING ARTIFICIAL INTELLIGENCE BUT THE OTHER ONE IS TO LOOK AT SPECIFIC MOLECULE WHERE IN THIS CASE, IN THIS LOCATION WE ACQUIRE THE SPECIMEN, ACQUIRE SPECTRUM AND THEN WE CAN ISOLATE SOME MOLECULES OF INTEREST, DO FRAGMENTATION AND IDENTIFY WHAT THE MOLECULE IS. THIS IS A LOW MASS ACCURACY INSTRUMENT, SO WE DO NEED TO DO TANDEM ANALYSIS TO BE ABLE TO DO THIS. BUT NOW, EVEN THOUGH THIS IS A LANGUAGE THAT WE SPEAK, IT DIDN'T MAKE ANY SENSE TO MY CLINICIAN COLLEAGUES SO THEN WHAT WE DID IS WE WORKED FROM SOFTWARE THAT WAS DEVELOPED AT BRIGHAM, SLICER WHICH YOU CAN DOWNLOAD AND IT'S USED TO VISUALIZE 3D DATA AND WE DEVELOPED MODULES WHERE WE CAN VISUALIZE NOW THE DISTRIBUTION OF THE TUMOR CELL CONCENTRATION BASED ON THE HISTOPATHOLOGY BUT ALSO VISUALIZE ANY MOLECULE OF INTEREST HERE. WE CAN MAKE THOSE LITTLE SPHERES TO SOMEWHAT TO SCALE AS TO THE SPECIMEN THAT WE TOOK HERE AND I'M NOT SEEING--THIS IS AN INTERESTING BIOMARKER BUT JUST TO SHOW THIS IS THE TOOL THAT WE'RE WORKING WITH TO BE ABLE TO COMMUNICATE OUR INFORMATION. SO, WE PUBLISHED QUITE A BIT ON USING ARTIFICIAL INTELLIGENCE, MACHINE LEARNING AND ALL OF THAT, BUT THE MOST MATURE PROTOCOL IS THE ONE WHERE WE FOCUS ON THE SPECIFIC METABOLITE TO SUPPORT SURGICAL DECISION MAKING IN THE BRAIN, SO IT'S IN THE W. H. O. PUBLISHED A NEW CLASSIFICATION OF BRAIN TUMORS OR THEY KEEP UPDATING THEM, AND WE'VE BEEN THINKING ABOUT THOSE TUMORS IN TERMS OF WHAT TYPE OF CELLS DO THEY LOOK LIKE, SO THEY'VE BEEN CALLED AN ASTRO SIGNIFY TOMA, DENDRITIC CELL ROW DPREEMOMA AND THEY'RE GRADED AND THIS IS A COMPLEX CLASSIFICATION SYSTEM WE'VE HAD BUT IN 2016, THEY REVIVED THAT CLASSIFICATION AND THE MOST IMPORTANT THING NOW IS TO FIGURE OUT IF THE PATIENT HAS A MUTATION OR NOT IN THE IDH. AND ALSO ISOTRAIT DEHIDE ROG NAIS, IT'S BECOME A POWER MARKER SINCE IF THE PATIENT HAS A MUTATION OR NOT, IT CAN INFORM THE WHOLE SURGICAL STRATEGY BASED ON THE FACT THAT THEY HAVE A DIFFERENT PROGNOSIS, DEPENDING ON IF THEY HAVE THAT MUSEUM TAIGDZ--MUTATION OR NOT. SO FOR THE CHEMIST OR MASS SPECT ROMMIVITY THIS IS WHAT IT LOOKS LIKE, SO THIS IS THE SPECTRUM FROM THE PATIENT WITH THE WILD TYPE ENWRIEM AND I HAVE TO SAY, OH I'M SKIPPING--SORRY, I SKIPPED SOMETHING HERE, SO ONE TECHNOLOGY TRANSFER I SHOULD SAY IS THAT DERIDE ROG NAIS WHEN IT'S MUTATED, IT PRODUCES METABOLITES THAT CAN NO LONGER GO INTO THE KREBS CYCLE AND IT ACCUMULATES AT MILLI MOLAR LEVELS SO THIS IS NOW WE CAN LOOK AT THE PRESENCE OF THE METABOLITE AND THIS IS ABSENT HERE FROM A WILD TYPE. WE SEE THE PIQ HERE IN A PATIENT WITH THE MUTATION, BUT AGAIN WE'RE WORKING WITH AN ION TRAP. THIS IS THE TYPE OF ACCURACY WE HAVE. SO TO MAKE SURE THAT THIS IS IN FACT, THE METABOLITE WE'RE LOOKING FOR. WE PERFORM TWO ROUNDS OF FRAGMENTATION AND THIS IS WHERE WE BRING THE SPECIFICITY. SO TO VALIDATE THIS WE WORK FROM A COLLECTION OF ABOUT 50 SPECIMENS, THE CURRENT DIAGNOSTIC TEST IS TO USE AN ANTIBODY THAT SPECIFICALLY RECOGNIZES, THE ARE--ADMINISTRATIVE GENERATEDDINE EFTHIMIOSA DINE MUTATION AND SO EVERY SPECIMEN WE HAD BEEN TOLD THE PATIENT HAD A MUTATION, WE WERE ABLE TO DETECT THE METABOLITE AND THERE WERE THREE SPECIMENS THAT WERE--THAT WE DETECTED THE METABOLITE THAT WE WERE TOLD WERE NEGATIVE, SO THIS ONE HERE, THE PATHOLOGIST WENT TO THE DIFFERENT BLOCK OF TISSUE AND FOUND THAT THE PATIENT HAD THE MUTATION AND THOSE TWO SAMPLES HERE REMAINED NEGATIVE, BUT ONE PERFORMING GENERATED ELTIC ANALYSIS WE FOUND THAT THIS PATIENT HAD AN RGENINE, TO GLUE MARIOUS SEEN IN THE PATIENT. SO ONE THING I SHOULD SAY IS THAT RECEIVING INFORMATION FROM THE IMMUNE O CHEMIST RADIOY STAIN CAN TAKE WEEKS FOR THE CLINICIAN TO RECEIVE, WHILE FOR US TO DETECT THE HYDROXY CLIEWTERATE, IT'S TO DETECT ONCE WE HAVE ISSUE, SO AT THE BEGINNING OF THE CASE, THE SURGEON GIVES US TISSUE AND WE CAN TELL IF THE PATIENT HAS A MUTATION OR NOT AND THEN THEY CAN--THAT'S ALREADY ONE ROUND OF SURGICAL DECISION MAKING THAT THEY CAN MAKE IS JUST TO KNOW WHETHER IT HAS A MUTATION OR NOT. SO WHERE WE PUSH THIS A LITTLE FURTHER, IS TO MAKE THE CORRELATION BETWEEN THE TUMOR CELL CONCENTRATION AND THE PRESENCE OF THE METABOLITE. SO HERE YOU CAN LOOK AT THE DISTRIBUTION OF THE METABOLITE AND THE INTENSITY OF THE METABOLITE CORRELATES VERY WELL WITH THE PERCENTAGE OF TUMOR CELL THAT YOU HAVE INTO THE TISSUE, SO NOW WE CAN, DOING THIS ANALYSIS ON THE SPECIMENS THAT THE SURMINGON IS INTERESTED IN, PROVIDE INFORMATION AS TO HOW MUCH TUMOR CELL BURDEN THERE IS IN A SPECIFIC PIECE OF TISSUE. AND THAT WORKS WELL FOR BRAIN TUMORS AS IT'S BASICALLY THE SURGEON PECKS AT THE TUBINGOR. IT'S NOT WHERE YOU GO AND MAKE AN INCISION AROUND THE TUMOR AND TAKE THE WHOLE THING OUT WHICH IS IN BLOCK RESECTION WHICH WE HAVE IN BREAST CANCER. SO IN THIS CASE, TAKING ALL THOSE TINY SPECIMEN WORKS WELL IN THE SENSE THIS IS HOW THEY DO THEIR SURGERY. SO NOW THIS IS THE FIRST REALTIME APPLICATION OF THIS, WHERE JUST TO GIVE YOU AN EXAMPLE OF THE WORK FLOW, SO WE HAVE IN THIS CASE, THE YELLOW--THE GREEN REGION IS WHERE THE TUMOR IS BASED ON THE PREOPERATIVE MR. AND IN YELLOW WE HAD THE LANGUAGE PATHOLOGISTSIA BASED ON MRI AND THE SURGEON WANTED TO REMOVE AS MUCH OF THE TUMOR AS POSSIBLE BUT PRESERVING THE HEALTHY REGION. SO WE DID ALL OF THOSE ANALYSIS HERE, AND WE'RE ABLE TO DETECT THE METABOLITE PROVIDED THAT INFORMATION AND WHEN THE SURGEON THINKS THAT MOST OF THE TUMOR IS GONE, OR IT BECOMES MORE CRITICAL TO KNOW WHAT THEY'RE DEALING WITH IN THAT OPERATING ROOM, WE CAN DO INTEROPERATIVE MRI AND THIS IS WHAT IS LEFT, THE TURQUOISE REGION HERE. SO WHAT HE DID WAS TO USE THE INFORMATION OF THE MASS SPECTROMETRY, ANALYSIS, TO DECREASE THE LEVEL OF THE METABOLITE BUT WITHOUT ACHIEVING ZERO IN THIS CASE. AND IT IS VERY IMPORTANT TO REMOVE AS MUCH OF THE TUMOR AS POSSIBLE, BUT IT'S AS IMPORTANT TO PRESERVE THE HEALTHY PART OF THE BRAIN. YES? >> [INDISCERNIBLE]. >> NO, SO WE TAKE THE LITTLE PIECE OF TISSUE, EXACTLY, SO THAT'S WHAT I WAS SAYING. THAT'S HOW THEY PROCEED, LIKE THEY TAKE A LITTLE PIECE AND THEY MOVE IT ASIDE, AND THEN THE ABILITY TO DO THE ANALYSIS THIS WAY HAS MADE IT SO THAT EVERY LITTLE PIECE OF TISSUE THAT WE GET WE CAN SPLIT IT INT TWO WHERE WE DO THE VALENTINEDATION AND THE ANALYSIS. BUT NOW WE CAN NO LONGER DO THIS FOR THIS PROTOCOL BECAUSE IT TAKES US TOO LONG. LIKE SHE WANTS TO KNOW RIGHT AWAY, SO AS SOON AS SHE GIVE ITS TO US, WE DO THE ANALYSIS BUT WE'VE DONE THE VALIDATION SO MUCH. SO UNTIL THERE, WHAT WE WERE ABLE TO TELL THE SURGEON IS WELL, THERE'S MORE OF THE METABOLITE--EXCUSE ME IN THIS SPECIMEN THAN THERE WAS TWO SPECIMENS AGO, OR IT WAS QUANTITATIVE. SO WHAT WE'VE DONE IS TO MAKE THE ANALYSIS QUANTITATIVE. SO WE PREPARE A TISSUE HEM EDIC WHICH IS BASED ON A COLLEAGUE PUBLISHING THIS FOR DRUG ANALYSIS AT GSK AND WE TAKE A GELATIN MOLD AND WE CREATE HOLES WHERE WE MAKE HUMAN EXTRACT FROM THE HUMAN BRAIN AND WE SPIKE WITH NONCONCENTRATION OF THE METABOLITE SO BEFORE A CASE, WE TAKE A CROSS SECTION OF THIS, MOUNT IT ON THE SLIDE AND WE CAN BUILD A STANDARD CURVE FROM THIS AND COMPARE HOW MUCH IS IN THE NEW SPECIMENS USING THAT STANDARD CURVE BUT THE OTHER COMPONENT WAS WE ADDED TO THIS WAS TO USE A TOPIC LABELINGEDICAL BRANDT WHERE WE HAVE CORRELATION CO EFFICIENTS ABOVE .99. SO USING THIS NOW, WE CAN TELL THE SURGEON, SO THE FROZEN SECTION IS WHEN THEY SEND A SPECIMEN TO THE PATHOLOGY ROOM AND WANT TO KNOW, IS THIS TUMOR NOT TUMOR, WHAT MIGHT IT BE, THEN WE WERE ABLE TO TELL HOW MUCH OF THE METABOLITE THERE IS, WHICH ARE DIAGNOSTICALLY RELEVANT CONCENTRATIONS HERE, BUT THEN AS THE SURGEON MOVES ALONG THIS SURGERY AT THE DIFFERENT MARGINS, THEN WE PROVIDE THOSE NUMBERS AND START TO DECIDE, YOU KNOW WHAT TO DO WITH THE INFORMATION. SO SO FAR, BASICALLY WHAT WE'RE WORKING TO DO NOW IS TO BRING SOME OF THE ROBUSTNESS AND STRINCHENCY THAT YOU HAVE IN CLINICAL PATHOLOGY, WHICH IS THE PART OF PATHOLOGY THAT ANALYZES URINE SPECIMENS, BLOOD SPECIMENS AND IN THIS CASE THE DIAGNOSTIC TESTS, IF YOU LOOK AT TEST RESULTS, IT'S BASICALLY THERE'S A RANGE OF WHAT IS NORMAL, ABNORMAL AND YOU REACH A THRESHOLD AND NOW ABOVE THAT IT'S ABNORMAL. AN ATOMIC PATHOLOGY IS THE ONE WHERE THE PATHOLOGIST HAS TO LOOK THEA THE TISSUE IS IT CAN BE VERY SUBJECTIVE AND VERY MUCH GRAY ZONES, NOW WHAT WE'RE TRYING TO DO IS TO BRING SOME OF THE SORT OF THRESHOLD QUANTITATIVE ANALYSIS TO THE ANALYSIS OF TISSUE TO HELP MAKE THOSE DECISIONS. AND EVEN THOUGH WE'RE USING MRI, WE'VE BEEN USING MRI AND OTHER TYPE OF IMAGING TO VALIDATE THE MASS SPECTROMETRY, NOW WE CAN GO AROUND FULL CIRCLE WHERE WE CAN--WE MAP THE MOLECULAR INFORMATION BACK INTO THE MR SPACE AS IT RELATES TO MR SEQUENCES WITH QUANTITATIVE INFORMATION AND TO TRY TO BETTER UNDERSTAND WHICH SEQUENCE OR WHICH COMBINATION OF MR SEQUENCE IS THE MOST TELLING OF TRULY WHERE THE TUMOR IS. SO NOW THIS IS WHERE IF I SWITCH GEARS TO SHOW YOU A LITTLE BIT OF WORK ON PROTEIN, WE DEVELOPED SOME APPLICATIONS FOR PITUE TARY TUMORS, SO THE TUMORS, THE MICROADNEMMA ARE SMALL TUMORS THAT ARE OFTEN RADIO LOGICALLY SILENT BUT THEN THE PATIENTS SHOW UP TO THE CLINIC WITH SYMPTOMS INDICATING THEY MIGHT HAVE A TUMOR ON THE PITUE TARY AND THE WAY THE SURGEON CAN MAKE THEIR STRATEGY IS TO FIGURE OUT BY SINUS SAMPLING WHICH SIDE MIGHT HAVE A HIGHER ELEVATION OF HORMONE LEVEL AND GO AND SACRIFICE THAT HALF OF THE TUMOR. SO OBVIOUSLY A TOOL THAT WOULD BE ABLE TO BETTER DELINEATE WHERE THE TUMOR IS, WOULD BE EFFECTIVE AND IT'S EASY TO UNDERSTAND. YOU WANT TO REMOVE THE TUMOR BUT AGAIN, YOU WANT TO PRESERVE THE SECRETING FUNCTION OF THE NORMAL PITUE TARY FOR THE PATIENT. SO IN THIS CASE, WE ARE DOING ANALYSIS, WHERE THIS IS ACTUALLY WHEN WE WERE PLANNING THE THE AMIGO, WHERE THE MASS SPECTROMETER IS IN THE ROOM, I SHOULD HAVE SAID LIKE THE USE OF AMBIENN IONIZATION, IT DOESN'T WORK REALLY WELL FOR PROTEIN ANALYSIS, SO WE USE A DIFFERENT TOOL FOR THIS, AND NOW, WE'VE PERFORMED THIS AND THIS SORT OF WE'RE WORKING TO BRING THE MALD I INSTRUMENT IN HERE. THE WHICH I WILL SHOW YOU WHAT IT IS SO WE CAN DO SOMEWHAT OF A FROZEN SECTION ANALYSIS BUT RAPID WITH WITH MOLECULAR INFORMATION. SO THIS IS THE LIST OF HORMONES WE HAVE BEEN TASKED TO LOOK INTO IN BOLD ARE THE ONES WE'VE BEEN ABLE TO ANALYZE USING MALD I IMAGING AND BASICALLY MALD I, MASS SPEC ROMETRY IMAGING STANDS FOR MATRIX LASER ARBSOPGZ IONIZATION SO IN THIS CASE, THE PREPARATION IS TIME CONSUMING.& WE TAKE A FROZEN TISSUE SECTION, APPLY THE MATRIX WHICH IS A SMALL ORGANIC ACID THAT CRYSTALLIZES ON THE SURFACE OF THE TISSUE AND SERVES TO TRANSFER THE ENERGY COMING FROM A LASER TO THE ANALYTES IN THE TISSUE. WE CAN ROSTER DELAYS IN AN X Y FASHION OVER THE TISSUE AND EVERY POSITION OF ANALYSIS BECOMES A PIXEL WHICH HAS AN ASSOCIATED MASS SPECTRUM OF A RANGE OF INTEREST AND THEN WE MINE THE DAILY BASIS AT DIFFERENT WAYS AGAIN. WE CAN VISUALIZE SPECIFIC MOLECULES FOR EXAMPLE, MOLECULE GREEN HERE PURPLE OR YELLOW. SO TYPICALLY IT WILL TAKE ABOUT AN HOUR TO DO GENERALLY THIS TYPE OF ANALYSIS AND A BIG TIME CONSUMING STEP IS THE MATRIX APPLICATION AND THE INSTRUMENTS THAT ARE BEING USED TO DO THAT, SO WE'RE SUBMITTING A MANUSCRIPT NOW WHERE--IN THIS MANUSCRIPT--NOT HERE, IN THIS PIT TUE TARY STUDY HERE, IT WAS TAKING US DOWN--WE WERE ABLE TO DO IT IN SOMETHING LIKE 30 MINUTES AND NOW WE'RE SUBMITTING A MANUSCRIPT WHERE WE CAN DO A MASS SPECTROMETRY IMAGING ANALYSIS IN LESS THAN FIVE MINUTES. SO THIS IS THE PROTOCOL NOW THAT WE'RE WORKING TO BRING INTO THE MOLECULAR FROZEN SECTION ANALYSIS AND THE REASON WE'RE ABLE TO DO THIS, ONE OF THE COMPONENTS IS A NEW INSTRUMENT FROM ONE OF THE VENDORS THAT IS MUCH MORE RAPID, BUT WE'VE ALSO BYPASSED SOME OF THE SAMPLE PREPARATION ISSUE AND LOADING OF THE SAMPLES, SO WE'RE PRETTY EXCITED ABOUT THIS. SO THE PITUITYARY TURNS OUT TO BE A NICE ORGAN TO WORK ON IN TERMS OF THE PROTEIN ANALYSIS AND WE KNOW WHAT THE PROTEINS ARE AND THEY HAVE VERY DISTINCT REGIONS WHERE EACH OF THESE HORMONES ARE BEING PRODUCED SO THIS IS HUMAN PITUEITARY WITH THE POSTERIOR PART AND THOSE HORMONES HERE WHICH ARE IN THE REGIONS WHERE WE WOULD EXPECT THEM, AND SOME THOSE WERE STANDARDS TO VALIDATE THEM IN THE HIGHER MASS BUT WHAT WE'VE DONE ALSO IS TO USE IN-SOURCE DECAY, AND IN THIS CASE WE USE A DIFFERENT MATRIX HERE AND WE'RE ABLE TO FRAGMENT THE PEPTIDE OR PROTEIN INTO THE SOURCE OF THE INSTRUMENT AND THEN PROVIDE PROVIDE SEQUENCE INFORMATION TO BE ABLE TO AND PROTEIN OF INTEREST. SO AGAIN, SIMILAR, AND ADENOMA THAT PRODUCE CERTAIN AND EXAMPLE HERE WHERE WE HAVE AND THERE'S ALSO ACTH HERE, SO NOW TO SEE IF THIS IS NORMAL ACTH, OR ABNORMAL SECRETION OF ACTH, WE CAN'T DETECT DISTINGUISHED THE TUMOR VERY WELL WITHIN HNE, AND THAT'S A STAINING THAT'S BEING DONE, TURNS OUT THIS IS THE NORMAL REGION OF THIS SPECIMEN AND THIS TAKES A VERY LENGTHY TYPE OF STAINING SO THAT'S NOT AVAILABLE DURING SURGERY. SO NOW THE QUESTION REMAINS THIS ONE WHERE I WAS JUST TALKING ABOUT THE CLINICAL PATHOLOGY VERSUS AN ATOMIC PATHOLOGY WHICH THIS IS WHAT THIS IS, IS HOW MUCH ECTH IS ABNORMAL. AND SO, ONE STRATEGY WOULD BE TO GO AND QUANTIFY AND RETERM WHRA THE THRESHOLD SHOULD BE IN A GIVEN REGION TO CALL IT NORMAL VERSUS ABNORMAL BUT NOW WE'VE COMBINED THE USE OF MACHINE& LEARNING WITH THE TARGETED ANALYSIS OF SPECIFIC PROTEIN AND FOR EXAMPLE, HERE, WE WERE ABLE TO CLASSIFY THE NONSECRETING VERSUS THE HORMONE COMING FROM A NONSECRETING VERSUS THE NORMAL VARY NICELY AND THIS NOW WE CAN GROUP--THIS IS JUST A VISUALIZATION USING PRINCIPAL COMPONENT ANALYSIS, SO THESE ARE MANY SPECIMENS OF ACTH SECRETING ADENOMA IF THIS IS GROWTH HORMONE SECRETING, PROLACTIC ACIDOSEISING VERSUS NONSECRETING AND THEY ALL SEPARATE NICELY DESPITE THE FACT THAT SOME OF THOSE SPECIMENS LIKE I'VE JUST SHOWN YOU MIGHT HAVE THE ACTH IN THERE. SO THIS IS ONE STRATEGY AND DOING THIS, WE SIGNIFICANTLY INCREASED THE SENSITIVITY AND THE SPECIFICITY OF THE ANALYSIS AND IN THIS CASE FOR EXAMPLE, HERE OF PROLECTIN WE HAVE GREAT SPEC PII SYSTEMS FISCHERITY BUT THE--SPECIFICITY BUT THE SENSITIVITY IS LOWER AS IT IS A VERY LARGE PROTEIN TO DEECTOMYOSIN ECTOMYOSIN INTO THE TISSUE--DETECT INTO THE TISSUE. SO NOW I WILL SWITCH GEAR AND GO THROUGH THE WORK WE'VE DONE ON DRUG DISTRIBUTION AND RESPONSE, THE MAIN ISSUE, WELL THE BLOOD BRAIN BARRIER IS A PHYSIOLOGICAL BARRIER THAT PROTECTS THE BRAIN FROM SURGULATING TOXINS AND PATHOGENS WHEN IT COMES TIME TO TREAT THE BRAIN, MOST OF THE DRUGS REMAIN WITHIN THE LUMEN OF THE BLOOD VESSELS AND GET WASHED OUT WITHOUT ACCESSING THE CANCER CELLS. SO WHAT WE WANT TO KNOW IS WHETHER THE DRUG IS FOUND WITHIN THE LUMEN OF A VESSEL VERSUS IF IT CAN ACCESS THE CANCER CELL AND IN THIS CASE, WE USE AGAIN MALD I MASS SPECTROMETRY IMAGING AND WE VALIDATE HEME, WHICH IS A FACTOR OF THE HEMOGLOBIN AS A MARKER OF THE VASCULATURE AND WE DID FLUORESCENCE IMAGING OF MULTIPLE SECTIONS OF ANIMALS THAT WERE--WE DID TAIL INJECTION OF FLUORA SEEN WHICH IS AN ACCEPTED MARKER OF THE VASCULATURE AND THE SAME TISSUE SECTIONS WE CODED WITH THE MATRIX DID MASS SPEC IMAGING OF HEME BUT ALSO A FLUORA SEEN AND SAW A STRONG CORRELATION. SO NOW IN THOSE STUDIES WE CAN IMAGE THE DRUG WITH THE MASS SPECTROMETER AND I DON'T THINK I NEED TO TELL YOU WE DON'T NEED TO PUT PROBES ON THOSE LABELS BECAUSE WE WORK FROM ACCURATE MASS, AND WE USE FTICR IN THIS CASE, AND OH, WE DOUBLE CHECKED IT BUT THERE'S DATA MISSING ON THIS. SO THE STORY GOES IN DRUG SCREENING FOR THOSE TUMORS IS THAT BASICALLY WHEN WE--NOW I HAVE TO--SO, WHEN WE PUT A BRAIN TUMOR IN THE PLANK OF AN ANIMAL, WE HAVE MANY DRUGS WHERE BASICALLY WE SEE A VERY STRONG EFFECT. BUT THEN THE PLACEBO GOES UP LIKE THIS, IT INCREASES IN SIZE, BUT WE HAVE A VERY STRONG EFFECT FROM A LOT OF THOSE TARGETED THERAPY. NOW IF WE PUT THE EXACT SAME TUMOR INTO THE BRAIN OF THE ANIMAL, WE LOSE THE EFFICACY, AND IT HAS BEEN OUR THOUGHT THAT THE BLOOD BRAIN BARRIER IS ONE OF THE BIG ISSUE AND NOW IT'S TO TRY TO UNDERSTAND--UNDERSTAND IT BETTER AND WHAT MIGHT GOVERN A DRUG FROM GOING, NOT GOING BECAUSE THE PREDICTING TOOLS ARE NOT THAT ACCURATE RIGHT NOW. IN ANOTHER COMPONENT THAT I WANTED TO SHOW HERE EMPLOY WHILE THIS IS A MOUSE BRAIN WITH THE TUMOR HERE AND THE WAY TO IMAGE THOSE TUMORS IS TO DO A CONTRAST MR AND INTO THE TISSUE, THIS IS WHERE THE IT IS THOUGHT THAT BASICALLY THE CONTRAST CORRELATESS TO THE BEST OF OUR KNOWLEDGE BUT YOU CAN SEE HERE THAT YOU HAVE A LOT OF THE CONTRAST THAT IS FOUND OUTSIDE OF THE TUMOR AND WE CORRELATED OF THIS DRUG HERE WHICH ACTUALLY WAS FOUND INTO THE TUMOR AND AND THOSE COMPOUNDS HAVE VERY SIMILAR MASS, BUT HAVE ALSO VERY DIFFERENT CHEMISTRY SO WE'VE DONE ENOUGH OF THOSE STUDIES WHERE WE WERE AWARDED BY THE NIH TO DEVELOP A CENTER TO DEVELOP IN THIS VERY AMBITIOUS TO DEVELOP PREDICTIVE MR BASED MODELS FOR CNS FOR THE FARM O KINETICS AND DYNAMICS, SO BASICALLY WOULD WE BE ABLE TO LOOK AT MR AND PREDICT WHERE A DRUG MIGHT GO AND HOW IT MIGHT EFFECT THE TISSUE. SO THS IS A CENTER FROM--THAT IS MIT IN COLLABORATION WITH MAYO CLINIC AND MY LAB, AND THE TYPICAL ANALYSIS IS THAT IT WILL GROW A TUMOR IN THE BRAIN OF THE ANIMAL AND PERFORM MRI, AND EACH MR PLANE WE ACQUIRE MULTIPLE TISSUE SECTION WHERE WE DO DIFFERENT TYPE ANALYSIS OF THE DRUG, VASCULATURE WE ALSO PERTUSSIS FORM SIMULATED ROMMENS MICROSCOPY ISSUING TO LOOK AT ARCHITECT AND YOU ARE LOOK AT TUMOR RESPONSE FROM THE PROTEOMICS FROM THE FLANK OF THE ANIMAL EVEN THOUGH DR. WHITE'S LAB IS WORKING TO DO IT FROM THE TUMOR ITSELF AND MRNA SEQUENCING THAT LINKS THIS AND DO VALIDATION WITH IMMUNE O HISTOCHEMISTRY, SO NOW IF WE WANT TO LOOK AT THE DISTRIBUTION OF THE DRUG IN THREE DIMENSION TO CORRELATE TO THE MR, WE TAKE CEREAL SECTION THROUGHOUT THE BRAIN OF AN ANIMAL AND TYPICALLY I SHOULD HAVE SAID TYPICALLY WE USE GREEN TO DEPICT THE BLOOD, AND NOW YOU CAN LOOK AT THE DISTRIBUTION OF THE DRUG ITSELF AND NOW CAN YOU LOOK AT THE DISTRIBUTION OF ONE OF THE METABOLITE OF THE DRUG AND RECONSTRUCT ALL OF THAT INFORMATION IN THREE DIMENSION HERE WHERE WE CAN LOOK AT THE VASCULATURE, BIOMARKER OF INTEREST, AND WE CAN SEE HERE, THE DISTRIBUTION OF THE PARENT DRUG WHICH IS TEN TIMES HIGHER THAN ONE OF ITS METABOLITE, BUT WE CAN SEE HERE THAT WE HAVE A VERY DIFFERENT DISTRIBUTION BETWEEN THE PARENT DRUG AND ONE OF THE METABOLITE AND THAT'S DESPITE THE FACT THAT THEY ONLY DIFFER BY A METHYL GROUP. SO THIS THE KIND OF INFORMATION NOW THAT WE CAN BRING TO FEED INTO OUR MODELS IN TERMS OF EVENTUALLY WHAT WE WOULD LIKE TO BE ABLE TO DO IS LINK PHYSICAL CHEMICAL IMPORTANTS OF A COMPOUND IN BETTER BE ABLE TO PREDICT ITS DISTRIBUTION. SO WE'VE ALSO MADE THOSE ANALYSIS QUANTITATIVE SO SIMILAR TO WHAT I'VE SHOWN YOU INTEROPERATIVELY. WE MAKE A TISSUE MEDIC FOR EVERY DRUG OF INTEREST AND NOW THIS IS WHAT IT LOOKS LIKE. IS YOU CAN QUANTITATE ONCE YOU'RE HAVE THE IMAGE, WE CAN QUANTITATE THE WHOLE DRUG IN THE BRAIN WHICH IS THIS NUMBER IN RED OR FURTHER SUGMENTORSHIP SKILL THAT THEY'VE DONE LOOK AT THE CONCENTRATION IN THE TUMOR ITSELF WHICH IS OVER TWO FOLD AND WE CAN ONCE WE HAVE ACQUIRED THE DATA, NOW WE CAN FURGTDER DISSECT A NORMAL REAMINGION VERSUS TWO DIFFERENT REGION WITHIN THE SAME TUMOR AND WE SEE A TWO FOLD DIFFERENCE WITHIN THE TUMOR ITSELF. SO NOW TO BE ABLE TO BRING THIS INFORMATION INTO THE MR SPACE, ONE OF THE CHALLENGE IS THAT THE MASS SPECTROMETRY IMAGING DATA IS ANY MOLECULE--IT'S BIOMOLECULAR INFORMATION. SO ONE MOLECULE DOES NOT GIVE YOU THE WHOLE INFORMATION OF THE SECTION AND WE WANT TO CORRELATE THIS WITH THE MRI, WHICH IS AN ATOMICAL INFORMATION. SO TO REDUCE THE DIMENSIONALITY OF THE THAT DATA WE USE TSME WHICH WE CAN MAP THE RESULTS ON TO THE BRAIN HERE, AND IT PULLS OUT SOME OF THE ANATOMICAL FEATURE THAT NOW WE CAN USE TO DO NONLINEAR REGITRATION TO THE MR AND NOW WE CAN, USING THIS, INFUSE THE MASS SPECTROMETRY DATA INTO THE MRI, AND NOW WE CAN SEE THE CONCENTRATION, HIGHER CONCENTRATION OF THE DRUG IN THE CORE OF THE TUMOR BUT WE DON'T SEE SO MUCH DRUG AT THE EDGE, AT THE LEADING EDGE--AT THE LEADING EDGE OF THE TUMOR. SO NOW TO BETTER UNDERSTAND WHAT'S HAPPENING INTO THE TISSUE, WE CAN USE IMAGING HERE IN THIS CASE, IN GREEN YOU HAVE THE GENERAL SIGNAL OF LIPIDS, IN BLUE TUMOR, IN RED THIS IS ABSORPTION TO LOOK AT THE BLOOD BUT WE CAN SEE THE CORE OF THE TUMOR AND IT WILL GIVE YOU THIS IS OPTICAL RESOLUTION SO YOU CAN SEE TUMOR CELL INFILTRATION AT REALLY HIGH SPATIAL RESOLUTION, THE DO NUT SHAPE OF THE RED BLOD CELLS HERE AND THEN WHEN WE OVERLAY THIS WE WERE ABLE TO SEE THAT WE, IF WE CORRELATE THE IMAGING WITH THE MASS SPECTROMETRY DATA WE CAN SEE THAT YES THE DRUG IS MOSTLY AT THE CORE OF THE TUMOR BUT WHAT WE SAW IS THAT WE HAVE WHITE MATTER PUSHING OR THE TUMOR FURBING AGAINST WHITE MATTER EFFECT ON DISTRIBUTION OF THE DRUG. AGAIN, I DON'T HAVE AN EXPLANATION FOR THIS, BUT THIS IS INFORMATION THAT WE'RE STARTING TO EXTRACT. SO, THERE'S THE ABILITY OF THE DRUG TO GO OUTSIDE OF A VESSEL BUT ONCE IT'S OUTSIDE OF THE VESSEL, IT'S ABILITY TO DIFFUSE OR TO BE TAKEN UP BY THE CANCER CELLS. SO NOW TO LOOK IN THE TUMOR RESPONSE, WE CAN USE THOSE APPROACHES, SO HERE IS A SUMMARY. THIS IS DONE FOR A LAB GROUP AT MIT AND THE MAYO CLINIC, SO IF WE LOOK AT THE TUMOR RESPONSE USING THE TRANSCRIPTOMICS HERE WE DON'T SEE MUCH OF A RESPONSE BUT WHEN WE'RE LOOKING INTO THE SIGNALING WE CAN SEE THAT IN FACT THE DRUG HAS HAD AN EFFECT ON THE PATHWAY THAT WAS TARGETED. THEN, IN THE SPATIAL RESOLUTION, WE CAN USE THE IMMUNE O EFTHIMIOS LOW CHEMIST RADIOY WHICH IS VALIDATED SO THIS IS A FOZ FOCUSED ON EFR STAINING AND WHEN THE FLANK TUMOR WAS EXPOSED TO THE TRUG, WE CAN SEE A REDUCTION INTO THAT STAINING. AND THE REASON I'M BRINGING THIS UP IS THAT FOR BRAIN TUMORS WE HAVE REALLY NOT HAD MUCH SUCCESS NEAR DECADES AND WE KEEP RUNNING TRIALS AFTER TRIALS. WE FAIL AND THEN WE PUT THE DATA ASIDE AND THEN WE GO AFTER THE NEXT MORE EXCITING COMPOUND. SOME GROUPS ARE FINDING THAT BASICALLY THE TUMOR CELLS WHEN YOU TREAT THEM AT SUB[INDISCERNIBLE] LEVEL, YOU INDUCE RESISTANCE MECHANISMS THAT MAKE THE TUMOR CELLS WORSE THAN IF YOU HAD LEFT THEM ALONE TO THEMSELVES. SO, BASICALLY NOW, GOING BACK INTO THE DISH, YAN FOUND WHEN HE TREATED THE CELLS AT LOW CONCENTRATION OF THE COMPOUND HERE, WE SAW AN INCREASE AND SAW MIGRATION THAT IS WORSE THAN IF QUIEWR NOT TREATING THE CELLS HERE. SO WHAT WE WANT TO BE ABLE TO ACHIEVE IS TO TREAT ALL THE CANCER CELLS AT THERAPEUTIC DOSAGE BUT EFFECTIVELY WE'RE ALWAYS TREATING AT SUBTHERAPEUTIC LEVEL AND MAKING THOSE TUMORS WORSE. THAT WAS NOT MEANT TO CHEER YOU UP AT ALL, IT'S ACTUALLY PRETTY DEPRESSING BUT HOPEFULLY WE CAN START UNDERSTANDING WHAT IS GOING ON AND TAKE A MORE RATIONAL APPROACH TO THE TREATMENT OF THOSE TUMORS. SO NOW EACH--HOW AM I DOING ON TIME? EACH OF THOSE SPECTRA FROM AN IMAGE IS FAIRLY COMPLEX, SO THIS IS ONE PIXEL FROM AN FTICR IMAGE WHERE THIS IS THE SIGNAL OF THE DRUG HERE, WHICH IS CAN ZOOM IN, THIS IS THE BLOOD SIGNAL, AND ALL OF THAT IS BASICALLY INFORMATION THAT WE THINK IS WORTH LOOKING INTO AS THEY CAN BECOME INTERESTING BIOMARKERS, SO THAT NOW WE COULD TRULY CORRELATE PK AND PD AT THE SINGLE PIXEL LEVEL. HOW MUCH DRUG IS THERE AND WHAT KIND OF AN EFFECT DOES IT HAVE? SO IF WE LOOK--AND OUR TRUSTEES IS BASED ON BEING ABLE TO DO THE TUMOR RESPONSE AND THE FLANK OF THE ANIMAL HERE AND EXTRAPOLATE THAT INFORMATION TO THE BRAIN, SO THESE ARE BRAIN WITH THE TUMORS HERE, SO WHEN WE TAKE ALL OF THAT IMAGING DATA AND WE SEGMENT THE DATA INTO TWO CLUSTERS, IT LOOKS LIKE THE FLANK TUMORS ARE VERY SIMILAR TO THE TUMOR FOUND IN THE BRAIN. AND/OR IT COULD BE HUMAN--HUMAN TUMOR VERSUS MOUSE DATA. BUT ONCE WE START REFINING OUR ANALYSIS AND ASK FOR EIGHT CLUSTERS, THEN THOSE ARE COMPLETELY DIFFERENT STRANGERS TO THOSE TUMORS HERE. AND THESE ARE AROUND THE TUMOR THAT WE DID FOR ANALYSIS OF THE DATA WE FOUND THAT BASICALLY A LOT OF THOSE MOLECULES WERE [INDISCERNIBLE], AND AND OF THE SALT TO AND THIS SHOWS THAT, AS A SOURCE OF ENERGY, SO NOW, TO TRY TO PATTERN, WE DID AGAIN LIKE THE 3D WITH THE MAPS SO THE AREA AROUND THE TISSUE AND REMAIN FOR ALL OF THOSE AND IN DIFFERENT ANIMAL MODELS, MANY ANIMALS OF EACH OF THOSE AND BEEN USING AS A--NOT SO MUCH AS A STANDARD BUT BUT I TOLD YOU IT SEEMS LIKE THERE'S MISSING DRUG AT THE EDGE OF THE TUMOR HERE AND WHEN YOU OVERLAP THOSE TWO SIGNAL IN FACT, THERE'S BASICALLY--IT SEEMS LIKE THERE'S EXCLUSION OF THE DRUG FROM THIS REGION AND IT TURNS OUT THAT ERLATNIB IS A DRUG THAT'S SOLLIBILITY IS VERY SENSITIVE TO PH SO WHAT WE'RE THINKING MIGHT BE HAPPENING HERE IS THAT METABOLISM--THOSE TWO DIFFERENT METABOLISM REGIONS HERE EFFECT DRUG DISTRIBUTION. THIS WAS JUST ACCEPTED AFTER, WORKING ON THIS APPLICATION OF THAT ANALYSIS FOR CLINICAL TRIALS SO THIS IS A PEYE KINASE INHIBITOR AND THIS IS THE MRI FROM THE PATIENT, THIS IS THE TIME OF THE FIRST SURGERY WHEN THE TUE TUMOR REOCCURRED, IT WAS TREATED FOR EIGHT DAYS WITH A HUNDRED MILLIGRAMS OF THE DRUG AND AT THE TIME AT THE TIME OF SURGERY, THE SURGEON TOOK REGION HERE AND NOW WE CAN LOOK AT THE DETAILED ANALYSIS OF ONE OF THOSE SPECIMENS AND BASICALLY AS I MENTIONED WHEN WE THINK CONTRAST AND HERE, AND WHEN WE LOOKED AT THE SPECIMEN, THE PATHOLOGIST DELINEATED THOSE WIGLY REGIONS HERE ARE NEUROECTODERMALCROSEIS AND THIS IS WHERE THE TUMOR HERE IS AND THIS IS ALL NORMAL TISSUE HERE, BUT IN THE BILK OF THE DRUG IN THIS CASE, WAS FOUND IN THE NORMAL PART OF THE TISSUE. SOPHISTICATED IF WE HAD TO JUST KINDLE SPECIMENS, IT'S ONLY IN THE PAST FEW YEARS WE TRY THESE TYPES OF TRIALS WHERE WE RECEIVE A DRUG PRIOR TO SURGERY BUT TYPICALLY THOSE SPECIMENS WOULD BE SENT FOR LCMS ANALYSIS ASK WE WOULD SAY, WELL, THE CONTRAST REGION WAS ALL VIABLE TUMOR AND THIS IS HOW MUCH DRUG THERE IS BUT WITHOUT THAT TYPE OF COMPARTMENTALIZATION HERE AS TO WHAT IS REALLYGOING ON. AND WHAT WE WANT TO BE ABLE TO DO IS TO CORRELATE THIS TO RESPOND SO THIS IS A SPECIMEN FROM THE PATIENT AT THE TIME OF THE FIRST SURGERY. THIS IS ALL SOLID TUMOR AND THESE ARE JUST SOME OF THE MOLECULES FOUND IN THOSE SPECTRA THAT HAVE HUNDREDS TO THOUSANDS OF SIGNAL AND IF WE COMPARE TO SECOND SURGERY, THERE ARE MOLECULES THAT ARE JUST COMPLETELY DIFFERENT. AND OF COURSE, IT'S NOT TO SAY, THIS IS THE BEST MARKER EVER, A LOT OF THINGS HAPPEN WITHIN EIGHT MONTHS AND THERE'S OTHER--THERE ARE OTHER TREATMENTS THAT ARE INVOLVED LIKE THE STANDARD OF CARE, BUT THIS IS DEFINITELY A PLACE TO START. SO WHAT WE'RE WORKING TOWARDS, DOING IS TO INCLUDE THOSE TYPES OF ANALYSIS TO AS MANY CLINICAL TRIALS AS WE CAN, SO WE CAN GAIN AN UNDERSTANDING OF THE DRUG DISTRIBUTION AS IT RELATES TO THE TISSUE ARCHITECTURE BUT ALSO IN THE 3D ANAL ME OF THE TUMOR AND AS IT RELATES TO THE 3D ANATOMY OF THE BRAIN WHICH AGAIN I DON'T THINK I MENTIONED THAT, BUT THE BLOOD BRAIN BARRIER IS NOT--HOMOGENERATEDDOUSLY REGULATED THROUGHOUT THE BRAIN, SO THERE ARE REGIONS THAT ARE MORE OR LESS PERMISSIVE AND THOSE ARE THINGS THAT WE'RE BRINGING SOME OF--HOPEFULLY CONTRIBUTING INFORMATION INTO THE SENSE OF BASICALLY YOU COULD START THINKING ABOUT TREATING A TUMOR IN A REGION VERSUS ANOTHER REGION, YOU MIGHT BE ABLE TO ADJUST A FARM CO--DOSAGE TO TAKE INTO,A KOWBT TO WHERE THE TUMOR IS. SO IN SUMMARY, I'VE SHOWN YOU MOLECULAR IMAGING OR BASICALLY MASS SPECTROMETRY IMAGING APPLICATIONS FOR INTEROPERATIVE ANALYSIS OF SURGICAL MARGINS AND PROVIDE APPLICATIONS EDUCATIONAL DIAGNOSTIC INFORMATION IN REALTIME, A LOT OF WORK THAT WE'RE DOING NOW IS TO ELECTRIC AT DRUG DISTRIBUTION AND RESPONSE WITH CORRELATION TO TISSUE CHARACTERISTICS AND BETTER UNDERSTANDING WHAT GOVERNS DRUG DISTRIBUTION AND RESPONSE IN WHAT WE'RE DOING IS CLINICAL TRIALS. SO I'M THE ONE WHO'S HONORED TO BE HERE AND TALK ABOUT THIS WORK BUT THIS IS EXTREMELY COLLABORATIVE WORK. SO I'M SURE AGAIN, I TRIED TO REVIEW THIS HERE AND THERE BUT I'M SURE I AM FORGETTING SOME PEOPLE, BUT IT'S VERY MUCH OF A TEAM EFFORT TO DO THIS. AND I WOULD BE HAPPY TO TAKE QUESTIONS. I THINK I DID THIS ON TIME, RIGHT? >> THANK YOU! [APPLAUSE ] I AM SLIGHTLY OUT OF BREATH, I BEFORE I CAME IN, BUT--YEAH? >> [INDISCERNIBLE]. >> WE DON'T. WE DON'T DO IT, THE FASTER PROTEOMICS ARE DONE ON THE FLANK TUMORS AND FOREST GROUP AT MIT WHERE ALEXANDRA TRAINED. >> YES. >> YEAH. >> AND THEN NOW, OUR CROSS CORRELATION AS TO WHAT MIGHT HAPPEN SPATIALLY IS WITH IMMUNE O HISTOCHEMISTRY AND THE KEEP R NA SEQ, THE SECTION OF THE DIFFERENT REGIONS OF THE TUMOR BASED ON HOW MUCH DRUG IS THERE AND SEE WHAT KIND OF CORRELATION WE HAVE, BUT AS FAR AS THIS IS NOT IN TERMS OF AND TUMOR FROM THE BRAIN SO THAT EVENTUALLY WE CAN DO IT, THE FOSTER NURSED FOCUSED ON PROTEOMICS FROM THE IRPT GRAIN KRAL TUMOR DOES THAT MAKE SENSE? >> YEAH. >> SO THERE'S WORKING ON PROTOCOLS, BUT IT'S NOT IMAGING. >> [INDISCERNIBLE]. >> YEAH, YEAH, I DON'T THINK--WE HAVEN'T TRIED BECAUSE WE'VE BEEN SO BOWSY WITH EVERYTHING. EVERYTHING ELSE, BUT YOU CAN DO PROTEIN ANALYSIS, SO THE ONES THAT I'VE SHOWN YOU I'VE SHOWN YOU THERE THAT WE'RE AFTER SPECIFIC HORMONES AND IT'S A VERY CLEAN TISSUE BECAUSE THOSE HORMONES ARE FOUND AT SUCH HIGH LEVELS BUT CAN YOU ALSO DO TRANSCRIPTIC DIGEST OF THE TISSUE INSITUE TRANSCRIPTIC DIGEST AND THEN DO THE MASS SPECULATION AND CONCLUSION TIDE IMAGING, AND THERE ARE DIFFERENT STRATEGIES TO DO IT AND THAT'S NOT ONE I WOULD DO ON THE DIAGNOSTIC TIME SCALE. AND THEN AND THEN FOR THE DRUG STUDIES, THEN FOREST IS THE BEST AT DOING THIS. SO I'M NOT GOING TO TRY. >> [INDISCERNIBLE]. >> YEAH, THAT WAS THE FIRST PAPER. THAT WAS THE FIRST PAPER, DOING BASICALLY YEAH, INSITUE PROTEOMICS. >> YEAH. >> [INDISCERNIBLE]. >> WE HAVEN'T BECAUSE I LIKE TO SAY WE WORK UNDER PEER PRESSURE IN THE SENSE THAT WE DEVELOP APPLICATIONS BASED ON WHAT THE CLINICAL NEED IS AND AND FOR THIS APPLICATION NOW, OUR NEXT STEP IS TO MAKE IT DURING SURGERY AND TO BRING THE INSTRUMENT THERE AND ALL OF THAT, RATHER THAN GOING THE OTHER ROUTE OF DOING ALL KINDS OF PROTEIN ANALYSIS, I MEAN THE DOCTOR'S GROUP HAS BUILT VERY MUCH FOCUSES ON THE ANALYSIS OF PROTEIN IN THE TISSUE. >> [INDISCERNIBLE]. >> YEAH. >> [INDISCERNIBLE]. >> OH YOU MEAN A PHOSPHOR O PROTEIN? >> YEAH [INDISCERNIBLE]. >> SO THAT'S WHAT WE'RE WORKING TO DO RIGHT NOW. SO ON THE PROTEIN SIDE, WE HAVE--WELL, AGAIN, FOREST'S GROUP, YAN'S GROUP WHO DO MORE LIKE THE SYSTEM BIOLOGY, THEY KNOW. SO THEY KNOW WHAT CHANGES, WHAT GOES UP, GOES DOWN, DOWN STREAM AND ALL OF THAT STUFF. OUR CHALLENGE NOW AND WHAT WE'RE WORKING TO DO IS TO CORRELATE THE METABOLISM WITH THIS, SO WHAT KIND OF INY TAB O LIMP RESPONSE DO YOU SEE WHEN THOSE THINGS GO UP AND DOWN. AND SO THIS IS WHAT WE'RE TRYING TO SORT OUT. BUT THAT'S A LOT OF--A LOT OF WORK AND WHAT WE'RE SEEING, I DIDN'T SHOW, SO ONE THING THAT IS A LITTLE UNSETTLING IS FOR EXAMPLE, WHEN WE DOD IS ITS WE TRY--WE DO STUDIES WE TRY TO MATCH THOSE PATIENTS SO THOSE ARE ALL. SO THE DOCTOR AT MAYO KNOWS WHICH ONE HAS A CERTAIN GENO TYPE SO IF WE DO A STUDY WITH THE THERAPEUTICS OF INTEREST THEN WE MATCH THE TUMORS TO MATCH THE GENETIC BACKGROUND AND WE ASSUME THEY'RE COMPARABLE BUT WHEN YOU LOOK AT THEM, THIS ONE STUDY WHERE WE COMPARED THE THREE DIFFERENT PDX WHICH ARE SUPPOSED TO BE VERY SIMILAR, THEY'RE COMPLETELY DIFFERENT. COMPLETELY UNRELATED AT THE METABOLISM LEVEL. SO,--BUT THERE'S HOPE. BECAUSE I THINK WHEN THE DRUGS--WHEN THE TUMORS START RESPONDING, I THINK SOME OF THE FIRST SIGNALS THAT FOREST TELLS ME THEY SEE ARE METABOLIC ENZYMES THAT CHANGE. SOPHISTICATEDY EVEN IF WE COULD JUST HAVE A RESPONSE, NONRESPONSE, DEATH OR SOMETHING LIKE THAT, YEAH. AND THE TWO H G WAS A BIOMARKER THAT HAS BEEN IDENTIFIED BY OTHER GROUPS WHILE THE IDH MUTATION BASED ON GENETICS AND THEN WE STARTED EARLY AS IT WAS GROWING AND THEN IT BECAME THE MOST IMPORTANT THING WE'VE HAD FOR GLIOMA AND NOW WE HAVE A GOOD TOOL TO ASSESS IT. >> [INDISCERNIBLE]. >> AS FARs I KNOW WE'RE THE ONLY ONES. BUT NOT THAT IT NEEDS TO BE SO I WOULD LOOK INTO VENDORS BASICALLY TO DEVELOP THE PLATFORMS NOW, SO THAT IT CAN BE DISSEMINATED BECAUSE-- >> [INDISCERNIBLE] >> EXACTLY. MM-HMM. YEAH. DO I HAVE TIME? >> [INDISCERNIBLE]. >> OH, SO WE HAVE--I'M HAPPY TO SHOW YOU IMAGES AFTER, BUT BASICALLY WHAT WE DID IN THIS CASE IS WE TOOK A COLLECTION OF PROOF THE ECTOMYOSIN ME WHICH CONTAIN LESIONS OF DIFFERENT LESIONS AND SCORES, SIX, THREE + FOUR, FOUR + THREES AND NINEs AND WE CAN SEE CHANGES IN METABOLITES THAT CAN DISTINGUISH TUMOR FROM THEA SOMEWHAT NORMAL PART OF THE PROOF THE ECTOMYOSIN ME, THAT CHANGE REGARDLESS OF GRADE BUT WE CAN ALSO SEE SOME THAT ARE INCREASED IN THE HIGHER GLEESON SCORES COMPARED TO THE LOWER ONES, BUT I THINK MOST INTERESTINGLY, WE CAN SEE CHANGES IN WHAT IS THOUGHT TO BE THE NORMAL COMOPPOSITE BEHAVIORIAL PHENOTYPENT OF THE PROOF THE ECTOMYOSIN ME ADJACENT TO THE INDEX LESION AND THAT IS MORE PROMINENT IN A PROSTATES THAT HAVE A HIGHER GLEESON SCORE LESION TO THEM. SO BASICALLY IF YOU LOOK AT COMPLETE PLEA NORMAL PROSTRATE VERSUS A PROSTATE THAT CONTAINS A HIGH FLEECEON SCORE TUMOR, THE NORMAL IS VERY DIFFERENT. AND IT'S SEEN IN RADIOLOGY BY USING ADC VALUES, BUT IT SEEMS TO BE HOLDING THROUGH INTO THE--AT THE TISSUE LEVEL. AND NOW WE'VE DONE THIS SO EVERYTHING WE DO IN TERMS OF THE BIOMARKER, ANOTHER INTERESTING THING IS THAT ONE OF THE MOLECULES OR SOME OF THE MOLECULES THAT INCREASE IN THE HIGHER DPLEESON SCORES, ARE AGAIN THOSE CARNATIENS THAT SIGNIFY THAT THE RESULTS MIGHT BE UNDERGOING BETAOXIDATION. >> [INDISCERNIBLE]. >> [APPLAUSE ]