Period From 1/2019 Through 12/2019
The group held a conference calls on 1/18 (originally scheduled for 1/11), 3/15, 5/10, 6/21, 8/2, 9/27, and 12/5.
Jan. 18: Dr. Testerman reported that experiments are continuing in a good direction and she is seeing tumors in the mice she is working with. Dr. Testerman said that the tumors do not have the appearance of lymphomas. She has ordered an antibody testing kit to determine if the tumors have an epithelial origin. In some cases she has seen positive tumor markers, e.g. CEA, but she wants to be confident about any conclusions she draws about the tumors. Dr. Testerman further reported that a number of her in vitro cell lines are doing well. In one recent experiment she injected mucin into a mouse and she is waiting to see what results. Currently she has found no new bacterial strains and she has a few bacteria that still need to be cultured and analyzed. A new undergraduate student has been added to her lab and this student should be helpful in general and with in vitro experiments in particular.
Prior to the conference call Dr. Merrell emailed the group an extensive PowerPoint presentation written by his postdoc Faith on her microbiome research. In addition 3 Excel files with detailed results for each of the 3 microbiome kits used (DNeasy, PowerSoil, QiaAmp) were emailed as well. The research was aimed at sorting out potential contamination issues with the microbiome kits that Dr. Blum was using. It was suspected that the kits themselves could be contributing hits to the bacterial results that were being recorded. This problem could arise because of the low amount of bacterial DNA in the PMP specimens being studied. The contamination could come from human contact with the water, pipettes, tubes, and reagents used in the testing process. The water that was used was purchased as DNA free water, but it did show up as containing bacteria during the sequencing. In addition to testing PMP specimens, the kits themselves were tested as was the water that was used. A total of 6 PMP specimens were tested. The results for the DNeasy, PowerSoil kits were not good and were described as giving results that were messy. Essentially the results obtained for the kits themselves and the PMP specimens overlapped. The family level results for these two kits showed no patterns at all. Statistically the kits and the PMP specimens could not be distinguished from one another. By contrast, for the QiaAmp kit although there was some overlap between the PMP specimens and the kit itself the results were dissimilar. There was a statistically significant difference between the specimens and the kit. For the QiaAmp kit it may be possible to subtract out the kit contributions from the PMP specimen results. Additional detailed results can be found in the PowerPoint presentation. The conclusions reached at this stage of the work are: 1. For the DNeasy and PowerSoil kits the PMP samples, kits, and water are similar; 2. For the QiaAmp kit the PMP samples and kit are (statistically) dissimilar; 3. the contamination from the kits is overwhelming the low microbial biomass from the PMP samples. Drs. Blum and Merrell will follow up with Dr. Metcalf about the problems identified with the kits and get her input on how to proceed. Dr. Merrell also mentioned that a new paper was published in the last few days that dealt directly with the problem of kit contamination. A question was raised about how the present approach to measuring the microbiome was different from that used earlier by Dr. Merrell’s Ph.D. student, Jeremy. In his case the DNA was extracted in Dr. Dubois laboratory before Jeremy carried out his microbiome testing. Although there was no negative control used, ISH staining done by Dr. Semino-Mora agreed with what was found in Jeremy’s microbiome study.
Dr. Merrell mentioned that the opinion paper was submitted to the European Journal of Surgical Oncology on Nov. 16. He checked the paper status before today’s conference call and noted that the paper was out for review and the last date listed for a status update was Dec. 26. A discussion took place about how long this journal typically takes to complete a review. Ms. Sittig said that in some cases reviews are done in 1 month and in others 3 months, so we are right in the middle of this span. Also, the holiday period could be affecting the review process. Dr. Merrell will wait a couple of more weeks before contacting the journal.
Dr. Testerman agreed in the last conference call to send photos of the tumors she is seeing to Mercy. She has ordered a camera and will follow up when it arrives.
March 15: Dr. Testerman reported that she continues to see tumors in mice but in some recent cases the development of tumors was disappointing. One mouse injected with mucin has developed a large girth and she is hopeful that she will see tumor in this case. Dr. Testerman mentioned that the tumors she has seen look like lymphomas but staining had shown that they are not lymphomas. She also mentioned that mucin is easier to work with than solid tumor and that mucin has significant viable cells in it. Homogenizing tumor can result in the destruction of cells contained in the tumor. In recent work she has studied 3 different groups of mice. One group was injected with tumor plus a mixture of bacteria, a second group was injected with tumor plus a mixture of bacteria plus antibiotics, and a third group with tumor only. Dr. Testerman wants to compare the rate of tumor growth in the 3 cases. The amount of bacteria injected is significant and on the order of 106 organisms per injection. No peritonitis has been seen from the injections. Dr. Testerman reported that she has not found any new bacterial isolates since the last conference call. She is considering changing her experimental methodology to increase her success in finding isolates. Dr. Merrell mentioned that he gave a talk at the FDA and found out that they have a well-funded project on whole genomic sequencing of bacterial isolates. It may be possible for Dr. Testerman to have some of her isolates sequenced as part of this FDA study. Dr. Merrell asked Dr. Testerman to send him a list of isolates that she would like to have sequenced and he will pursue the matter with his contacts at the FDA. He suggested that the list be sent in the next week.
After the last conference call on Jan. 18 Dr. Merrell and Dr. Blum telephoned Dr. Metcalf to get her input on their microbiome work. They were interested in getting her input on the data from the 3 kits they tested as discussed in the last conference call. Dr. Metcalf has extensive microbiome experience. Although the results from the 3 kits studied looked different statistically, Dr. Metcalf said that because of the large differences in reads obtained from the kits, we should not worry so much about the statistics. For samples where smaller numbers of reads were obtained, achieving statistical significance can be difficult. Conversely, in instances where millions of reads were obtained, very small differences could result in statistical significance but might not mean anything biologically. Dr. Metcalf suggested that inputting more material to the kits should help. Dr. Merrell initially was reluctant to sacrifice all the material he has for some samples, as a result smaller amounts of material were used for the tests. After getting the results discussed in the last conference call, he has decided to input larger amounts of material to increase yield. Dr. McAvoy suggested that when the next analysis is done that the three ovarian cancer specimens that Mercy shipped to Dr. Merrell be tested. If we can develop results on ovarian cancer then our ability to generate funding should increase dramatically. Dr. Merrell also mentioned that he has been asked to sit on an NIH panel to evaluate a PMP proposal. He attributed this invitation to the publication of our latest paper on antiobiotics and PMP. A link to this paper can be found on the publications page of this website. The panel is an R21 panel associated with the NCI Clinical and Translational Exploratory/Developmental Studies. This area of NIH should be a good place to consider for submission of future PMP proposals.
Dr. Nieroda reported that 28 patients have been enrolled in the antibiotics protocol, and that 4 of these patients had stopped taking the antibiotics. Twenty two patients were enrolled in the fecal microbiome study and due to patient drop out there are now 9 antibiotics patients and 9 non- antibiotics patients left in this study. Dr. Testerman reported that she has sent pictures of the tumors that she has been observing in mice up to Mercy. She was encouraged by some of the similarity between the tumors she has been seeing and those seen in the PMP patients from whom the material was harvested and implanted into mice. Dr. Testerman also mentioned that for cases where she receives a large amount of tumor, she plans to do microbiome studies on the specimens. Dr. Merrell raised the issue about Mercy getting whole genome sequencing of tumors done. Ms. Sittig has developed a proposal for such sequencing and Dr. Merrell has pushed this proposal to the genome center at USUHS to see if there is if they have any interest in collaborating on this project.
May 10: Dr. Testerman reported that she has changed her experimental procedure to filter out host cells and she is seeing more bacteria as a result. Dr. Testerman also reported that the nude mice she has been experimenting with are heavy with a weight over 30 grams. Normally these mice are not that large and they are showing signs of tumor growth. Dr. Testerman and her technician are in the process of finishing up some papers and thus, they will have more time for the PMP project in general and in vitro work in particular. Dr. Testerman mentioned that she has become aware of information on the microbiome giving rise to ascites in ovarian cancer, and she plans to study this information.
Ms. Sittig reported that progress on recruiting additional antibiotics patients has been slow. Two patients required fast surgery and there was not enough time to start the antibiotics protocol. Thus, there have been no new patient enrollments. One new patient was enrolled in the fecal microbiome study and 3 to 4 patients need to complete the necessary questionnaires for the University of San Diego so that their specimens can be processed. Ms. Sittig said that she expects that a batch of specimens will have been sent to San Diego before the next conference call. Concerning control tissue, 2 to 3 additional specimens are still needed. Ms. Sittig said she would check into this issue.
There were two follow up items from the last conference call. Dr. Testerman sent bacterial isolates to Dr. Merrell so that they could be cultured by Jeanette and submitted to the FDA for genome sequencing. Dr. Merrell checked with the FDA and they do not have clearance to accept the live bacterial cultures. Therefore, the bacteria will be cultured in his lab and stored as frozen pellets. Hopefully these bacterial pellets will be sent out to the FDA next week. The second follow up item involved two proposals from Mercy Medical to the genome center at USUHS. One proposal involved sequencing PMP tumors, and the other sequencing ovarian cancer tumors. Dr. Merrell forwarded these two proposals to the center and they expressed an interest in the ovarian proposal, but not so much interest in the PMP proposal due to the fact that PMP is a rare disease. However, the center did not give up completely on the PMP proposal. Ms. Sittig mentioned that Dr. Sardi is trying to find private donors to fund the PMP proposal.
During the conference call a new paper1 on PMP from Mercy Medical was discussed, as well as a paper2 on a new drug that can be used to break up mucin. Immediately after the call Ms. Sittig emailed copies of both papers to the group along with a copy of Dr. Sardi’s PMP genome proposal.
New Papers: 1. Selection and Characteristics of Patients with Perintoneal Dissemination from Appendiceal Cancer with Exceptional/Poor Survival after CRS/HIPEC, Carlos Munoz-Zuluaga, et al, published online, Ann Surgical Oncology. 2019 Apr 30. doi: 10.1245/s10434-019-07374-z. 2. Assessment of a novel mucolytic solution for dissolving mucus in pseudomyxoma peritonei: an ex vivo and in vitro study, Pillai K, Akhter J, Morris DL, Pleura Peritoneum. 2017 Jun 1;2(2):111-117. doi: 10.1515/pp-2017-0013. Epub 2017 Jun 6.
June 21: Dr. Testerman reported that her experiments with mice continue to go well. In one experiment she injected PMP-derived tumor from one mouse into new mice. One half the mice were given an injection of PMP191F bacteria while the other half did not get the injection. PMP191F in vitro appears to slow down tumor growth. Interestingly, only the mice receiving the PMP191F bacteria developed tumors. Dr. Testerman speculated that perhaps the bacteria helps the tumor cells survive in vivo. Dr. Testerman is carrying out additional experiments with different bacteria and she is waiting to obtain results. Dr. Testerman mentioned that mice that have been injected with cell lines or tumors don’t start dying after intra peritoneal (i.p.) injections (of saline, bacteria, or antibiotics) until several months after tumor injection and when they are necropsied, she finds copious amounts of blood in the peritoneal cavity. Dr Testerman has performed i.p. injections on thousands of mice over the years and never had them die suddenly, so she thinks that the mice already have bloody ascites at the time they are injected and that the injection sets off a coagulation cascade that proves fatal. Dr. Testerman believes that this is similar to Trouseau’s syndrome, which is a coagulopathy that occurs in humans with mucinous tumors. It is believed that the mucin reacts with some clotting factors. Heparin is used in patients with this syndrome and she will now add heparin to the solutions she injects to prevent the clotting cascade from getting started. Dr. Nieroda asked whether the ascites were seen only in the intra-abdominal cavity. Dr. Merrell asked about the origin of the mice used in these experiments. A discussion of these questions followed. Dr. Testerman also reported that she has seen one mouse develop a colorectal tumor with mucinous ascites. She has also seen mucinous ascites develop in mice injected with patient mucin. Dr. Testerman mentioned that so far, all tumors and cell lines have had at least some cells positive for CEA and MUC2. Sometimes they are extensively positive and sometimes it’s just one part of the tumor. She has stained three cell lines with CDX2 and found them to be positive. The two cell lines that have been tested for CK-20 have been positive. The tumors Dr. Testerman has looked at so far just have a few scattered cells positive. This could relate to the tumor differentiation state. Poorly differentiated colon cancers are less likely to be CK-20 positive. Undifferentiated (mesenchymal) cells often express vimentin, so she will look for that. One undergrad is working in Dr. Testerman’s lab on immunohistochemistry and a medical student is working on in vitro studies. Dr. Testerman has also heard back from a female contact at a company that makes a kit to test 60 antibodies simultaneously. She will check the list of 60 to see which would be useful for the PMP research. Dr. Testerman noted that not every PMP patient specimen is useful for developing tumors and not every specimen produces a tumor in mice. Specimens from patient PMP 441, who happened to be a DPAM patient, have been very fruitful in developing tumors in mice. However, most of the tumors that develop in mice are solid tumors. Some tumors have mucinous ascites and many have bloody ascites. Dr. Sardi mentioned the genomic study that Mercy would like to undertake on PMP patients. He thought that it would be interesting to compare the genomes of patient tumors with those grown in mice. It would be interesting to see if a correlation exists between the pathologies of the two. Studying mice in this way might lead to a quicker knowledge of what to expect in patient outcomes. Dr. Testerman has studied numerous PMP patient specimens. She is in the process of putting her results into a spreadsheet. It was suggested that once completed the spreadsheet could be sent to Mercy for comment and comparison with patient records. Dr. Testerman agreed to follow up on this suggestion.
The one follow up item from the last conference call involved completing the paperwork for the fecal microbiome specimens to be analyzed at the University of San Diego. The majority of the questionnaires have been completed and only 2 to 3 remain to be completed. At present we have 19 microbiome fecal samples to be processed, 9 from patients receiving antibiotics and 10 from non-antibiotics patients. Ms. Sittig will coordinate with Dr. Metcalf when she gets back from maternity leave about submitting the specimens.
It was mentioned that Mercy is in contact with a few companies interested in the PMP genomic project and a proposal is pending with the ACPMP Research Foundation to fund this work. Dr. Merrell mentioned that the request to USUHS to sequence PMP isolates has not been turned down, and it is still being considered by USUHS for future work. Dr. Merrell also mentioned that the isolates that Dr. Testerman sent to him about 1 to 2 weeks ago have been forwarded to the FDA for sequencing.
Aug. 2: Dr. Testerman reported that another animal in her study was found dead in the cage. She believes that it had not been dead very long and when it was examined, it had visible ascites, what appeared to be mucinous tumor, a tumor in the liver and the liver was discolored. They recovered the ascites and trypan blue staining suggested that some of the cells were alive, so these were injected into another animal. The interesting thing about this animal was that it was originally injected with a PMP cell line that was derived from a patient (PMP457, DPAM patient). Cells from the patient tumor were cultured in vitro for a period of time to create the cell line and then these were what were injected into the animal approximately 1 year ago.
A very thin animal was also euthanized, but there was no obvious tumor. Additionally, a new bacterial isolate from a recent PMP tumor sample was cultured. This new isolate is filamentous and looks a lot like PMP215, though she believes it is different species. 16S sequencing will be performed next week for identification.
She submitted an abstract to the NORD conference. Mercy mentioned that they also submitted several to the same conference. Among these, Ms. King mentioned that one of their abstracts is focused on their recent analysis that indicates that the number of epithelial cells found in the mucus may negatively correlate with tumor progression; the more cells, the faster the tumor will progress.
Dr. Testerman said that she met someone from GE that has the ability to stain tissue slices with up to 60 antibodies; this is a new machine that they want to test. They have agreed to look at a few of the PMP samples and she is picking particular antibodies that will target pathways of interest (differentiation, cell signaling, Muc2, etc.)
Finally, she said that she is planning on two upcoming papers; a mouse model paper and an in vitro tissue culture model paper. Additionally, the original 191F paper is almost ready to be resubmitted.
Dr. Merrell mentioned that he had set up a collaboration for the sequencing of these samples. The plan is to sequence 96 samples (including sequencing controls) to start. If these work well, the number of samples will be expanded in subsequent rounds of sequencing. It was suggested by Dr. McAvoy that the ovarian cancer samples be included in this round. Mercy suggested that we not include the tumors from the interesting familial case. The hope is to start prepping samples in about 2 weeks.
Dr. Testerman’s PMP strains were cultured and sent to the FDA for sequencing.
Ms. King reported that one new patient was added to the microbiome study and one new patient was added to the antibiotic study since that last conference call. Current numbers for microbiome are 10 patients on antibiotics, 10 not on antibiotics and 4 withdrew; the goal is 24 patients total. Current totals for the antibiotic study are 20 enrolled and 9 withdrew. One new control specimen was sent to Dr. Merrell.
Ms. King mentioned that they also submitted the familial case to the NORD meeting as an abstract. They are developing an extensive data collection on this mother and daughter; germ line sequencing, appendiceal tumor sequencing, fecal microbiome, the tumor microbiome, etc.
The one follow up item from the last conference call involved Dr. Testerman sending a spreadsheet to Mercy on the samples that she has been processing and injecting into animals. She did this and Mercy is working on completing their portion of the spreadsheet.
Sept. 27: Dr. Testerman reported that she has not cultured any new isolates. Also, her funding for the mice has been exhausted, so any new animal research will await additional funding. As a result, Dr. Testerman has been euthanizing mice and, in some cases, she has seen abnormalities that appear to be tumors; she is waiting on results from pathology. Dr. Testerman has been freezing material for future animal research, which she feels will be easy to restart when more money for animals is available. In the case of some mice, Dr. Testerman has been able to re-isolate bacteria injected into mice. She has re-isolated PMP215, which is a waxy, branched bacterium. Interestingly, unlike other bacteria this bacterium decreases host cell respiration, and Dr. Testerman has been able to quantitate the decrease. PMP215 is similar to other bacteria that have been found in the PMP microbiome. Dr. Testerman has also re-isolated a Streptococcus species as well. A discussion about the bacteria sent to the FDA for sequencing took place. PMP215 was one of the species that the FDA will sequence. The specimens were sent to the FDA in late June and hopefully results should be available shortly. Ms. Sittig raised the point about making sure the material Dr. Testerman has implanted is actually positive for cancer. Species have been harvested at the time of surgery and sent to Dr. Testerman for implantation. Pathology results are obtained sometime later. Ms. Sittig said she would provide pathology information for the specimens sent to Dr. Testerman.
Dr. Merrell discussed his plan to sequence 96 microbiome samples. His Postdoc, Dr. Blum has selected the 96 samples as follows:
35 DPAM subjects (24 female, 11 male)
35 PMCA (mucinous adenocarcinoma) subjects (21 female, 14 male)
6 other cancer subjects (4 ovarian, 1 mucinous ovarian, 1 uterine carcinosarcoma)
6 PMP CONTROL samples (peritoneal tissue)
Controls: 8 no-input controls; 6 positive controls (dilution series of mock community)
The no-input controls are used to detect reagent contamination. The specimens to be analyzed were selected based on suggestions that were received from the group. In choosing samples, priority was given to specimens that had a majority of tumor since these produce more DNA than mucinous specimens. The size of the microbiome specimens is 250 mg, and if there is not enough tumor then the specimen has mucin added to it. The DNA preparation for PCR should be completed today. Carrying out the PCR reactions should take 1 to 2 weeks. The samples produced by PCR will be sent to a collaborator at BDRD, which is a military facility in Frederick Maryland. Analyzing the microbiome of the 96 specimens should take about 1 month. Once the results are available Dr. Merrell said he would email them to the group. Dr. Merrell briefly discussed our recent antibiotics PMP paper and noted its publication. Dr. McAvoy checked on Researchgate.net and found that the paper had 27 readers.
Ms. Sittig reported that there were no new fecal microbiome patients enrolled since the last call. Ms. Sittig noted that we need 5 additional patients to make the 24 that are sought. We currently have 10 abx and 9 non-abx patients in the study. Ms. Sittig mentioned that 1 new surgical abx patient was enrolled giving a total of 21 patients in the surgical study. The surgical abx study is close to 3 years old and it will need to be renewed with Mercy’s IRB. Ms. Sittig did not feel that there would be a problem with the renewal. Ms. Sittig said she would email data on the 21 patients, such as diagnosis, status, etc. to the group before the next conference call.
Dec. 5: Prior to the conference call Dr. Testerman emailed copies of pathology slides from her mouse studies. Dr. Testerman had discussed the slides yesterday with a pathologist at her institution. A detailed discussion of what Dr. Testerman’s slides showed took place. Dr. Testerman mentioned that even when a slide did not contain tumor in many cases it was not totally normal. One of her slides contained signet ring tumor from a specimen from a patient with signet ring disease. In one of the mice the liver showed a great deal of tumor and bleeding. This result could explain the occurrence of bloody ascites in some mice. Other mice exhibited large pancreases, which led to a question about whether PMP patients might be hyper-glycemic as a result of the disease. Dr. Gushchin stated that hyperglycemia is not seen clinically with PMP patients. Another mouse exhibited tumor on the outside of the spleen. An enlarged spleen indicates irritation. Dr. Testerman mentioned that she has had success using peritoneal lavage to get solutions from which cells can be cultured. Even when mice show no signs of tumor per se but they simply look sickly and are losing weight, peritoneal lavage has allowed Dr. Testerman to culture cells. Dr. Testerman mentioned that her slides have contained a mixture of tumor types, with both low and highly malignant tumors showing up in the same mouse. Dr. Testerman’s results correlate with disease type, either DPAM or PMCA. Dr. Studeman will compare Dr. Testerman’s slides with slides from the patients from whom Dr. Testerman’s specimens were obtained. Dr. Testerman has tried using antibiotics on mice to see if they help with disease progression but whether the antibiotics have helped remains to be evaluated. Dr. McAvoy suggested that Dr. Testerman probably has enough material to write a couple of papers, which could be co-authored with people from Mercy Medical. Dr. Gushchin asked what hypothesis would be tested in each paper. Dr. Testerman said that one paper could focus on her animal model and compare her mouse results with patient tumor pathology. A second paper could focus on new methods of culturing cells in vitro. Dr. Testerman has been successful in culturing cells from both DPAM and PMCA patients and her culturing results could be correlated to a patient’s clinical outcome. Ms. King mentioned that for DPAM some patients can do poorly and it would be interesting if culturing results could shed some light on why. Lastly a discussion about organoids took place.
Dr. Merrell updated the microbiome research. The 96 specimens that his postdoc Faith chose were successfully amplified. These specimens were sent to the sequencing facility and Dr. Merrell expects to have results back before the holidays. At that time he will be able to compare the PMP microbiomes with those of controls. Dr. Merrell mentioned that Faith will be leaving on Jan. 3 to take up an IRTA Fellowship at NIH. He will need to identify someone to continue the work that Faith has been doing. Dr. Merrell also updated the status of the PMP isolates that Dr. Testerman had sent him to be sequenced at the FDA. Roughly 1/3 of the isolates have been sent on for sequencing and results should be available reasonably soon from the FDA. Included in the specimens sent on were bacteria from PMP patients 215 and 238. Dr. Testerman mentioned that the PMP215 results would be particularly interesting because of the type of bacteria present.
Ms. King reported on the various studies underway at Mercy. After the call she emailed the information below about these studies. For the fecal microbiome project 24 people were enrolled but 5 were off study. Five more patients are needed to get to the goal of 24. Of the 19 patients enrolled, 10 have taken antibiotics and 9 are non-antibiotic patients. For the PMP antibiotics study one new patient was enrolled but that patient withdrew because of going to another institution. Ms. King also provide an overview of all the PMP patients at Mercy. Since 9/27/2019 14 new patients were enrolled. Specimens have been collected on 12 of them. Overall 261 specimens have been collected from 335 enrolled patients. Control specimens from 25 patients have been collected. Recently the sample size for collecting PMP specimens was increased from 350 to 1000 through Mercy’s IRB.
Prior to the conference call Ms. King mailed out two very detailed documents. Both were discussed during the conference call. The Antibiotics Study Update was a 17 page Research Summary giving results from 11/4/2014 to 10/18/2019 for our PMP antibiotics study. Patients in the study take pre and post-operative antibiotics in conjunction with cytoreductive surgery and HIPEC. The following information, taken from Ms. King’s report, was discussed. Of the 217 patients screened for inclusion in the antibiotics study, only 30 were enrolled and currently 19 remain in the study, but results for 18 were included since the last patient has not yet started preop antibiotics. Numerous reasons exist for patients failing the screening including such things as allergies to antibiotics, advanced disease, patients declining enrollment, not enough time before surgery, negative malignancy, among others. Patients who went off the abx protocol did so for reasons such as surgery was aborted, patient went to another institution, aborted HIPEC, or palliative HIPEC. For the patients remaining in the abx study 89% completed the preop antibiotics regimen, but only 50% the postop regimen. Patient refusal is the primary reason for the postop percent being low. The cytoreductive surgery often leads to patients struggling with diarrhea, nausea, and vomiting, and patients do not want to add antibiotics. Of the 18 antibiotics patients 12 were DPAM, 3 PMCA, 2 PMCA signet cell, and 1 PMCA goblet cell. Of the 18 patients 13 had a CC (completeness of cytoreduction) score of 0 and 5 a score of 1. At present all 18 patients are surviving and only 1 has experienced disease recurrence. Significant additional data is given in Ms. King’s report. The second document discussed was a tumor tracking pathology specimen excel spreadsheet. The spreadsheet gives details on the pathology reports from PMP patients at Mercy and it also details the specimens that were sent to Dr. Testerman for her culturing and animal studies. This information will be very useful in comparing Dr. Testerman’s results with what was found clinically. The group complimented Ms. King on the thoroughness of her work on the two reports.