 So in my presentation, I'm going to share with you the methodology to monitor the drug efficacy that we use in our project in Cambodia, the malaria project. So it's a second presentation on Cambodia. I make the presentation on behalf also of the colleagues that are listed on the screen. Now, first, a few words on the project. So the overall objective of the project is to develop strategies to eliminate the plasmodium fulciparum from that area of Artemisinin resistance. So you know Artemisinin is the most potent drug we have against malaria, definitely an indispensable drug. So the reason why we are working on malaria in Cambodia is specifically to tackle that resistance. Now, it's obvious, it's common sense that the last resisting parasites and the last strains are going to be the most resistant ones, which means that if we want to tackle resistance, ultimately we will have to work towards elimination in such an area. So the reason that we are there is of course to avoid the spread of the resistance from that area to other areas. So on the map you see the place where we are working, so the district Chaisan bordering Thailand and you see the study period. We started in October 2015 and I have data until April 2018. So within that broad objective of the project we have a number of specific issues, questions linked to the efficacy or the resistance if you want. So the first one is more on the public health level. We want to make sure that if there would be a decrease in the efficacy of the ACTs, so the Artemisinin based combination therapy that we are using, that we would be able to detect this in an early stage. We need this to know obviously because if this would be the case we will have to change the drugs or eventually the drug regimen. Or if it would not be black-white this can be then an indication to do a full-scale full-blown efficacy study which is very heavy because this means we have to do PCR, we have to do microscopy and for a single patient we need at least 10 samples, day 1, day 2, day 3 and so on up till day 42, so which is heavy for the patient and quite resource intensive. And then on the individual level we want also to detect in an early stage if someone is not responding well to a treatment because if someone is not responding very likely to be resistant, a resistant parasite, we don't want that this person continues to contribute to further transmission. Now what we do concretely in the project is we do of course case finding which means we test and treat patients with fever. We do reactive case finding which means that if we have a positive malaria case we consider it as an index case and we test around, that's the household and people working in the similar places where probably the transmission is ongoing. And we do proactive case finding which means we do send MSF staff to the villages where they test people on a regular basis, people that have risk exposure, concretely working in the forest. We also have basic vector control and so we do the monitoring of resistance in vitro. I come back to that. So in fact where does the question come from though? We have in the project a strengthened case management which is of course not the routine way of working that we have in Congo or in Africa but we have a strengthened case management system not for research purposes but simply we want to make sure that in that area with resistance a patient is treated and the parasites are indeed eliminated. And so the question that came up was can we not if we use know that opportunity of a strengthened case management if we would know structure and standardize the follow-up could we not use this and evaluate this as a methodology, a lighter methodology to monitor the drug efficacy or the resistance if you want. So let me first explain now what it means that strengthened monitoring of the patients as we do it. So if we have a case, usually rapid diagnostic test, we do systematically take at day zero a dry blood spot that is analyzed in the central lab in Phnom Penh, the Pasteur Institute lab. So the dry blood spot that we take and then we give all the patients a three days directly observed treatment because we cannot in such a situation work with incomplete treatments or accept incomplete treatments of course. And you see that we have been using two different ACTs in the first period, the hydro-artemisinin-piperaquin and because of the resistance we had to switch later on to artesionate mefloquin. Then for the follow-up of the patients we go back, we trace back the patients, all the patients at day 28 and we take again a dry blood spot. We get the result normally within a week and so we want to make sure that indeed the patient has eliminated all the parasites. Then specifically in the frame of a study we added also day 63, I will come back to that, but this was to validate if day 28 was indeed a good time and a good indicator for us to base our methodology on. And then there's the further monitoring that takes place mainly in the laboratory of Pasteur where they follow up on our in vivo samples and that's then for the treatment failures, the distinction between re-infection, recrudescence. They also calculate the estimated paracetamia based on the PCR as we don't have microscopy and they follow the different molecular markers that are correlating with resistance and so that's the ones correlating with artemislin resistance, mefloquin and piparquin resistance. Now the three specific questions to which I will come back one by one is first of all, we want to know are our ACT still efficacious, is day 28 a good choice and if we make such an analysis should we include all the patients. Now for the first question, are the ACTs that we have been using, are they still efficacious enough? And so what we did here, what you see is the PCR based paracetamological treatment failure on day 28. So if we look at the first row and maybe just look at the percentage and have the confidence interval as well, so we see that when we're still using DHAP peraquin, we had 36% treatment failures and even more concerning we know we saw that the vast majority were recrudescences, so meaning the parasites were not eliminated by the drug. Because of that resistance we had to switch to an alternative drug which is still today artesianate mefloquin and you see that the second row there we only have 7% treatment failures and moreover there is not a single recrudescence among these patients, it's all reinfections. I just have to mention that unfortunately we had quite some samples with insufficient genetic material to do DD analysis and it's based on relatively small numbers. So this first of all gave us a very clear conclusion, the use of dehydro-artimizin peraquin is definitely no longer justified in that area. The second question is, is day 28 an acceptable time to do that evaluation? The question comes from the fact that normally in a standard efficacy study we always do day 42. But we are using PCR, which means we are using a more sensitive technique and moreover we want to make sure that the patient not responding to treatment is identified quite early. And so you see that for that reason we did day 63 and so here you see and we only did it in the artesianate mefloquin period you see that from the 261 patients from whom we had day 28 negative that only 7% had day 63 positivity rate and moreover there was not a single case of recrudescence which for us led to the conclusion day 28 is indeed a good time to do it, it's not too early to base our conclusions on. The third question of the methodology was then should we include all patients and this question comes from the fact that in the standard efficacy studies normally we exclude all the patients with a parasitemia below 1000 parasites per microliter the reason is simply that if you start very low and if you take your drug that even in case of resistance you may not yet have sufficient parasites to be detectable during the follow-up period. So on this slide you see the same data on the left that I showed you already and we did the exercise over again excluding the samples where we had the parasitemia load below 1000 parasites and you see indeed looking again at the percentage that while in the initial one we had 36% treatment failures this becomes 59% treatment failures in the first row so with our DHAP Periquin with touching confidence intervals showing that indeed if we include the low parasitemic ones that we may underestimate the resistance or overestimate the efficacy as it's written on the slide. So the conclusions for us is that first of all the implementation is feasible it's of course not the routine data that we have in our African projects but in such a place where we want anyhow the patient to get 3 days directly observed treatment and we want anyhow to make sure that we have a follow-up after 28 days and we see that we managed to trace back at least 60% of the patients for that follow-up on the 28 so we see that the PCR based day 28 treatment is indeed an appropriate indicator it's not too early and we see that if we would include all the patients including the low parasitemic ones that we may overestimate the treatment efficacy so it does make sense to exclude these patients from the analysis. Now however have we been using these results or what was the relevance? First of all as we could document that high failure rate of the DHAP Periquin this clearly was a reason to say that we should change also in the area where we were working as a priority the ACT to the alternative one which was in this case the artesanate mefloquin and then as there were similar results from other areas this led at the end to the nationwide decision to drop the first line DHAP Periquin and change it everywhere to artesanate mefloquin and so we realize of course even if I did put in the title routine data this is not again the routine for our African setting but we think that this methodology of using what we have can be relevant in places where we do have first of all few cases which means pre-elimination or elimination setting so where we can afford a close monitoring of the individual patient and especially where we are confronted with a high level of resistance because that's the places where we cannot wait until these full scale efficacy studies are done which sometimes takes two, three years before we have the results but just to make sure that we have nearly real time at least basic information about the efficacy of the drugs that we are using and that we do this on top of the in vivo monitoring sorry in vitro monitoring based on the molecular markers so I would like to thank our study partners so that's the Pasteur Institute in Phnom Penh and the National Malaria Control Program C&M and also our MSF Cambodian staff because they are doing the main work in the fields and the population of that district where we have been working for several years now Chesan the representatives of the population and you for your attention thank you