acfalcao ● 20070917 00:34 (6013 d 03:11 ago) Posting: # 1082 Views: 33,232 

Dear All My name is Amilcar Falcão and I am professor at the University of Coimbra, Portugal. I am looking for help regarding the execution and interpretation of threeway crossover bioequivalence trials (for academic purposes). I would like to have an example (explicit) where a threeway crossover study is appropriately analysed. Namely, I would like to know what kind of ANOVA should be performed and how the 90%CI should be calculated for the different combinations of study formulations (i.e. A vs B and A vs C). Perhaps this is a naive question, but in fact I am trying to look for it in literature and I have seen different approaches always without the adopted statistical pathway. Therefore, I would like to be sure regarding the correct way to deal with. I strongly appreciate if you could help me about this matter. Thank you for your attention. Best regards; Amílcar Falcão Laboratory of Pharmacology Faculty of Pharmacy University of Coimbra Coimbra  Portugal 
Helmut ★★★ Vienna, Austria, 20070917 22:47 (6012 d 04:59 ago) @ acfalcao Posting: # 1088 Views: 31,196 

Dear Amilcar! ❝ I would like to have an example (explicit) where a threeway crossover study is appropriately analysed. You have to use a Williams’ design (three period, six sequences); the topic was covered in previous threads. A 6×3 design is needed in order to ‘extract’ two 2×2 tables, which are also balanced. Although the full 6×3 table will be used in the analysis of AUC and C_{max}, you will need these 2×2s for the nonparametric analysis of t_{max} (unfortunately there’s no confidence interval based nonparametric method available for more than two formulations/periods). The asterisks * denote pseudosequences and pseudoperiods, e.g. P1* means only that the treatment was given in a period prior to P2* – irrespective of the true study period:+++ > +++ and +++ A common mistake is to design the study as a set of 3×3 latin squares, which will lead (especially if the sample size is small and in the case of drop outs) to extremely imbalanced data sets: +++ > +++ and +++ ❝ Namely, I would like to know what kind of ANOVA should be performed and how the 90%CI should be calculated for the different combinations of study formulations (i.e. A vs B and A vs C). For the design see Chapter 10 of Chow SC, Liu Jp. Jones B, Kenward MG. Patterson S, Jones B. You may download zipped datasets and programs (SAS/S+) from CRC’s website. If you don’t have access to commercial software, S+ code will run with opensource R with minor modifications. Patterson/Jones give results in Table 4.12 (p. 105) with: T/R (% Test vs. Reference 1) ++++ T/S (% Test vs. Reference 2) ++++ WinNonlin 5.2 comes up with: T/R (% Test vs. Reference 1) ++++ T/S (% Test vs. Reference 2) ++++ Results are slightly different (although both WinNonlin and SAS use GLM – not ANOVA; implementation, rounding, etc. is different – and of course ‘proprietary information’ and not documented). I assume Patterson/Jones' results were produced by SAS; I will check the results from their S+ code the next days. ❝ Perhaps this is a naive question,... Not at all; little is published  and no worked examples at all. There’s another point which is a little bit tricky: multiplicity. If you are testing one test (A) against two references (B, C), any impression of ‘data dredging’ must be avoided, e.g., calculation 90% CIs of A/B and A/C  and only picking out the best getting an approval. Since in the EU the Innovator's product from any European country may be used as the reference, you may run into problems (A vs B is BE, whereas A vs C is not BE). It may be wise to use 95% CIs instead (overall Bonferronicorrected patient’s risk: α = 1–(1–0.05/k)^{k}, where k is the number of simultaneous comparisons). 95% CI should also be applied in testing for dose proportionality of three dose levels (or 96.67% for four levels). IMHO the only case where 90% CIs should be used is the comparison of two test formulations against one reference, and only one of them will be further used in the approval process. — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
acfalcao ● 20070918 01:02 (6012 d 02:44 ago) @ Helmut Posting: # 1089 Views: 29,945 

Dear Helmut Thank you very much for your help. The correct codification of "sequences/periods" is the key for this problem. Unfortunately the link to download the SAS files was not working. I hope it can be solve in a near future. Best regards; Amílcar Edit: Full quote removed. Please delete everything from the text of the original poster which is not necessary in understanding your answer; see also this post #5! [Helmut] 
Helmut ★★★ Vienna, Austria, 20070918 14:47 (6011 d 12:58 ago) @ acfalcao Posting: # 1092 Views: 29,939 

Dear Amilcar! ❝ Thank you very much for your help. The correct codification of "sequences/periods" is the key for this problem. Unfortunately the link to download the SAS files was not working. I hope it can be solve in a near future. Please try it again (it worked yesterday, and today as well): CRC’s website Having downloaded the file C5300.zip, unzip it keeping folder's structure; in folder [chapter4] you will find: Example4.5.sas (the SAS code) Example45.SSC (the S+ code) exam45.dat (the raw data for S+) As I get from the SAScode a mixed model (degrees of freedom according to Kenward) was used, whereas WinNonlin uses Satterthwaite's method. The S+ code does not calculate BE, but is used for the example’s figures only. Edit: Wrong link corrected in both posts. [Helmut] — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
Nirali ★ India, 20070921 08:49 (6008 d 18:57 ago) @ Helmut Posting: # 1106 Views: 29,804 

Dear Helmut, Which hypothesis Winnonlin will use while running BE for 2 test (T1 & T2) vs a reference (R)? Would it be "MEAN(T1)=MEAN(T2)=MEAN(R)"? In threeway crossover study, does we try to compare Equivalence of all three product together? Thanks & Regards, Nirali 
Helmut ★★★ Vienna, Austria, 20070921 15:04 (6008 d 12:41 ago) @ Nirali Posting: # 1109 Views: 29,848 

Dear Nirali, ❝ Which hypothesis Winnonlin will use while running BE for 2 test (T1 & T2) vs a reference (R)? ❝ Would it be "MEAN(T1)=MEAN(T2)=MEAN(R)"? ❝ In threeway crossover study, does we try to compare Equivalence of all three product together? Comparisons are based on paired differences; IMHO a simultaneous test of T_{1}=T_{2}=R is not possible. WinNonlin always uses one reference in a run, i.e., T_{1}=R (one test), and T_{2}=R (the second test). If you are interested in T_{1}=T_{2} you have to start a second run (defining T_{2} as the reference), and will obtain two tests (T_{1}=T_{2} and R=T_{2}) within. BTW, why are you interested in T_{1}=T_{2}? — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
Nirali ★ India, 20070925 10:05 (6004 d 17:41 ago) @ Helmut Posting: # 1127 Views: 29,763 

Dear Helmut, Actully, I was under impression that WinNonlin is using the hypo: "MEAN(Test1)=MEAN(Test2)=MEAN(Ref)". As I understood from your reply (A) suppose we have two REFERENCES and single TEST [R1, R2, T]for threeway comparison: if we select R1 as REFERENCE, WNL will calculate BE with hypo: (1) R1=T and (2) R1=R2. same way, (B) for two TEST and single REFERENCE [T1, T2, R]: if we select R as standard, hypo will be: (1) R=T1 and (2)R=T2 pls do correnct me if i am wrong to understand. Thanks & Regards, Nirali 
Helmut ★★★ Vienna, Austria, 20070925 15:15 (6004 d 12:31 ago) @ Nirali Posting: # 1132 Views: 30,048 

Dear Nirali! ❝ Actully, I was under impression that WinNonlin is using the hypo: ❝ "MEAN(Test1)=MEAN(Test2)=MEAN(Ref)". Again, we obtain only pairwise comparisons, not simultaneous ones. ❝ (A) suppose we have two REFERENCES and single TEST [R1, R2, T]for threeway comparison: if we select R1 as REFERENCE, WNL will calculate BE with hypo: ❝ (1) R1=T and (2) R1=R2. Not quite, but very close. If you run the BE wizard (R_{1}=reference), you will get (1a) T vs. R_{1}, and (1b) R_{2} vs. R_{1}. You have to run the BE wizard again (this time with R_{2} as reference), getting (2a) T vs. R_{2}, and (2b) R_{1} vs. R_{2}. In order to get comparisons of T=R_{1} and T=R_{2}, you have to run the wizard twice (results 1a and 2a), omitting 1b/2b. Of course PE and CL of 2b=1/1b. ❝ same way, ❝ (B) for two TEST and single REFERENCE [T1, T2, R]: if we select R as standard, hypo will be: (1) R=T1 and (2)R=T2 Yes this is correct; in this case only one run of the BE wizard is necessary, getting: (1a) T_{1} vs. R, and (1b) T_{2} vs. R. — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
Irene_I ☆ Indonesia, 20180607 13:09 (2096 d 14:37 ago) @ Helmut Posting: # 18861 Views: 21,942 

Dear Helmut, I'm currently looking for statistics methods for William Design in Bioequivalence Study and I found this thread. I tried to download the SAS code from the link you've attached, but apparently the link doesn't work for now. ❝ Please try it again (it worked yesterday, and today as well): CRC's website ❝ Having downloaded the file C5300.zip, unzip it keeping folder's structure; in folder [chapter4] you will find: ❝ Example4.5.sas (the SAS code) ❝ Example45.SSC (the S+ code) ❝ exam45.dat (the raw data for S+) I know that this topic was discussed years ago, but I think I really need to download these files for my further understanding about this topic. I would really appreciate if you could help me with this. Thank you. Best Regards, Irene I 
Helmut ★★★ Vienna, Austria, 20180607 15:04 (2096 d 12:42 ago) @ Irene_I Posting: # 18862 Views: 21,953 

Hi Irene, ❝ I know that this topic was discussed years ago, but I think I really need to download these files for my further understanding about this topic. I would really appreciate if you could help me with this. The files of the 2005 edition are gone. However, here is the link for the 2016 edition. — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
Irene_I ☆ Indonesia, 20180608 13:09 (2095 d 14:37 ago) @ Helmut Posting: # 18869 Views: 21,834 

Dear Helmut, Thanks a lot! It is really a great help to me.. Regards, Irene I 
Irene_I ☆ Indonesia, 20180612 11:21 (2091 d 16:25 ago) @ Helmut Posting: # 18886 Views: 21,820 

Dear Helmut, Could I ask you some questions about this example? After I tried to run the SAS code (Example 4.5), I got the CI, but there is only one Residual value. I guess I suppose to calculate the intrasubject CV from this value, right? So, My question is, Do we only get one Intrasubject CV from threecomparison of the treatment (TR, ST, SR) while we got three CI (TR, ST, SR)? I am looking forward for your help. Thank you very much. Best Regards, Irene I 
Helmut ★★★ Vienna, Austria, 20180612 14:42 (2091 d 13:03 ago) @ Irene_I Posting: # 18888 Views: 22,556 

Hi Irene, ❝ After I tried to run the SAS code (Example 4.5), I got the CI, but there is only one Residual value. I guess I suppose to calculate the intrasubject CV from this value, right? So, My question is, Do we only get one Intrasubject CV from threecomparison of the treatment (TR, ST, SR) while we got three CI (TR, ST, SR)? I’m not equipped with … Hence, my understanding of the code is limited. You are right, in the model you get only one (pooled) estimate of the variance. That’s not a good idea, since it will “work” only if intrasubject variances would be identical and the treatment differences at least very similar. Otherwise, the treatment estimates will be biased and the type I error is not controlled. See also this post. At the 2^{nd} Workshop of the Global Bioequivalence Harmonisation Initiative (Rockville, September 2016) Pina D’Angelo gave a presentation “Testing for Bioequivalence in Higher‐Order Crossover Designs: Two‐at‐a‐Time Principle Versus Pooled ANOVA” showing exactly that. Here some of her slides: Purpose
Introduction Statistical Concerns:
Simulated Data: Summary
Closing Remarks
If you are interested in all pairwise comparisons, generate three data sets (values or T&R, S&R, T&S, i.e., exclude all values of the respective other treatment S, T, R) whilst keeping the codes for sequence and period. This will give you data sets which represent an IBD (incomplete block design). Run the usual model on them. This approach is also recommended in the EMA’s BEGL: In studies with more than two treatment arms (e.g. a three period study including two references, one from EU and another from USA […], the analysis for each comparison should be conducted excluding the data from the treatments that are not relevant for the comparison in question. (my emphasis)Example 4.5 (Phoenix/WinNonlin 8, mixed effects, Satterthwaite’s degrees of freedom): PE (%) 90% CI (%) s²_{w} CV_{w} (%) Evidently variances are not identical. The ones of T/R are smaller than the ones of S/R. If we apply the pooled model the CIs of T/R will be wider than in the IBD model and the CIs of S/R narrower. BTW, I don’t understand what the purpose of the carry variable in Byron’s code is.— Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
Irene_I ☆ Indonesia, 20180613 07:02 (2090 d 20:44 ago) @ Helmut Posting: # 18896 Views: 21,900 

Dear Helmut, Thank you for your explanation. I am sure this will be helpful not only for me, but also others who looking for information about this topic too. Best Regards, Irene I 
Helmut ★★★ Vienna, Austria, 20180613 17:02 (2090 d 10:44 ago) @ Irene_I Posting: # 18898 Views: 21,691 

Hi Irene, ❝ Thank you for your explanation. You are welcome. A picture tells more than a thousand words (AUC only). The PE and its CI of T/R are similar. The small bias is caused by the fact that the average PE of S/R and T/S (\(\sqrt{141.09\times83.12}\)) is close to the T/R with 116.05. The other comparisons show negative biases. The CI of S/R is narrower in the pooled model and the CI of T/S is wider. We don’t want to go there. — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
d_labes ★★★ Berlin, Germany, 20180613 17:31 (2090 d 10:14 ago) @ Helmut Posting: # 18899 Views: 21,792 

Dear Helmut, ❝ BTW, I don’t understand what the purpose of the As far as I see it: It serves for nothing because it is not used except in the class statement of Proc Mixed .The class statement serves the purpose of declaring factors even if a variable is numeric. Nothing more. The carry variable seems to me a left over from old times where carryover was an issue.— Regards, Detlew 
ElAlumno ☆ 20190315 00:40 (1816 d 02:05 ago) @ Helmut Posting: # 20036 Views: 18,529 

Dear Helmut, In your old but stilluseful post (excerpt below), you describe generating pseudosequences and pseudoperiods when assessing ABE. ❝ A 6×3 design is needed in order to ‘extract’ two 2×2 tables, which are also balanced. Although the full 6×3 table will be used in the analysis of AUC and C_{max}, you will need these 2×2s for the nonparametric analysis of t_{max} (unfortunately there’s no confidence interval based nonparametric method available for more than two formulations/periods). The asterisks Your post preceded the EMA's Guideline on the Investigation of Bioequivalence by a few years, but it appears to agree with their stipulation: In studies with more than two treatment arms (e.g. a three period study including two references, one from EU and another from USA, or a four period study including test and reference in fed and fasted states), the analysis for each comparison should be conducted excluding the data from the treatments that are not relevant for the comparison in question. I completely understand the use of pseudosequences in this situation. But what about pseudoperiods? The EMA guidelines aren't too specific. On the one hand, if you wish to display your data with pseudosequences, you would want to use pseudoperiods too (so that the timing of treatments is consistent within pseudosequence). On the other hand, the ANOVA doesn't care whether your pseudosequences are logically consistent with your period. If there is no period effect, the difference between using period and pseudoperiod will be minimal (use a few more df in the ANOVA table with period than pseudoperiod). But what if there is a serious period effect? You would miss it (wrong sum of squares for period) by using pseudoperiods. Opinions? 
Helmut ★★★ Vienna, Austria, 20190315 01:47 (1816 d 00:59 ago) @ ElAlumno Posting: # 20037 Views: 18,819 

Hi ElAlumno (nice nick!), ❝ In your old but stilluseful post (excerpt below), you describe generating pseudosequences and pseudoperiods when assessing ABE. ❝ ❝ ❝ A 6×3 design is needed in order to ‘extract’ two 2×2 tables, [] ❝ ❝ Your post preceded the EMA's Guideline on the Investigation of Bioequivalence by a few years, but it appears to agree with their stipulation: […] Not quite. ❝ I completely understand the use of pseudosequences in this situation. But what about pseudoperiods? The EMA guidelines aren't too specific. The EMA means the ‘Two‐at‐a‐Time Principle’ (see above) and not my recoding (which I abandoned a good while ago as well). David Brown (MHRA, member the EMA’s Biostatistics Working Party) gave a clarification at the 3^{rd} EGA Symposium on Bioequivalence (London, June 2010). The Q&A document states: However, the treatment, groups, sequences and periods should have their original values maintained in the analysis, and not have the values modified. For example an observation made in period 3 should still be coded as period 3, not have the period changed to “2” because the results for that subject in one of the earlier periods has now be removed. (my emphasis)❝ On the one hand, if you wish to display your data with pseudosequences, you would want to use pseudoperiods too … Nope. Relevant slides: Guideline — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
ElAlumno ☆ 20190322 22:59 (1808 d 03:46 ago) @ Helmut Posting: # 20069 Views: 18,074 

Thank you Helmut for your quick and detailed response! I'm glad to hear that pseudoperiods have gone the way of the dodo. I agree with the EMA Biostatistics Working Party that it makes more sense (statistically) to maintain the original codings when leaving out data from irrelevant treatments (not to mention being less prone to human error than recoding). Also, I love their name; I bet the EMA Biostatistics Working Party could form a coalition government with the Slightly Silly Party, the Surprise Party, and the Rent Is Too Damn High Party. But I digress. I don't have WinNonlin, but I tried analyzing Patterson & Jones' Example 4.5 with the Two‐at‐a‐Time Principle using R 1.1.456. My results were similar to the results you posted above, but not as close as I would have expected. I will post the code & table below. Do you see anything I am doing wrong or any obvious explanation of the differences? I don't know how WinNonlin handles missing values, so I tried a few variations (subjects with complete cases in the full dataset, and subjects with complete cases in each of the 3 incompleteblock subsets), but the differences persisted.
# 3treatment design from Patterson & Jones (2017) This produced the following table:
Test PE LCI UCI CV 
Helmut ★★★ Vienna, Austria, 20190323 02:03 (1808 d 00:43 ago) @ ElAlumno Posting: # 20070 Views: 18,161 

Hi ElAlumno, ❝ I agree with the EMA Biostatistics Working Party […]. Also, I love their name; I bet the EMA Biostatistics Working Party could form a coalition government with the Slightly Silly Party, the Surprise Party, and the Rent Is Too Damn High Party. It is really existing though David Brown will be brexited (is this already a word?) unceremoniously. ❝ But I digress. That’s fine. Adds some spice to that dry stuff. ❝ […] I tried analyzing Patterson & Jones' Example 4.5 with the Two‐at‐a‐Time Principle using R 1.1.456. What? 1.1.456? 1.1.4 was the first one I used in 2002. Current is 3.5.3! ❝ My results were similar to the results you posted above, but not as close as I would have expected. I will post the code & table below. Do you see anything I am doing wrong or any obvious explanation of the differences? I don't know how WinNonlin handles missing values,… That’s not the point. The devil is in the details. I wrote above: ❝ ❝ ❝ ❝ Phoenix/WinNonlin 8, mixed effects Why? Well, Patterson & Jones analyzed the data with SAS Proc MIXED and I wanted to reproduce it in PHX/WNL (restricted maximum likelihood). OK, they used in SAS ddfm=KENWARDROGER ^{1} and yours truly Satterthwaite’s df.When I switched in PHX/WNL to a fixed effects model (i.e., like lm() in R and SAS Proc GLM ) I got exactly your results. Hence, your code is fine though you can streamline it (specify NAs and classes):
I would not use option(..., digits=4) cause IMHO, this may affect also the precision of internal calculations. Better to round yourself at the end, f.i.
So what do we have (T/R of AUC)? $$\small{\begin{array}{llccc}\hline \text{Method} & \text{Setup} & \text{PE} & \text{90% CI} & \text{CV (%)} \\ \hline \text{pooled} & \text{SAS Proc MIXED (KENWARDROGER)} & \text{116.2 } & \text{109.0 , 123.9 } & \text{21.3}^2 \\ & \text{SAS Proc MIXED (SATTERTHWAITE)} & 116.15 & \text{108.97, 123.81} & 21.20 \\ & \text{PHX/WNL REML (Satterthwaite)} & 116.15 & \text{108.97, 123.81} & 21.20 \\ & \text{R lmer() Satterthwaite} & 116.15 & \text{108.97, 123.81} & 21.20 \\ & \text{R lmer() KenwardRoger} & 116.15 & \text{108.97, 123.81} & 21.20 \\\hline & \text{PHX/WNL LM (all fixed)} & 116.24 & \text{109.04, 123.92} & 21.22 \\ & \text{R lm()} & 116.24 & \text{109.04, 123.92} & 21.22 \\ \hline \text{IBD} & \text{SAS Proc MIXED (SATTERTHWAITE)} & 116.05 & \text{108.92, 123.65} & 20.84 \\ & \text{PHX/WNL REML (Satterthwaite)} & 116.05 & \text{108.92, 123.65} & 20.84 \\\hline & \text{R lmer() Satterthwaite} & 116.05 & \text{108.97, 123.58} & 20.84 \\ & \text{R lmer() KenwardRoger} & 116.05 & \text{108.97, 123.58} & 20.84 \\\hline & \text{PHX/WNL LM (all fixed)} & 116.14 & \text{108.98, 123.78} & 20.88 \\ & \text{R lm()} & 116.14 & \text{108.98, 123.78} & 20.88 \\ \hline \end{array}}$$If Satterthwaite’s df are used, the mixed effects model in SAS and PHX/WNL agree. R lm() agrees with PHX/WNL’s fixed effects model. A little bit surprising to me that Proc MIXED with ddfm=KENWARDROGER^{3} gives the same result as a fixed effects model. OK, but the FDA recommends Satterthwaite’s df for ages.^{4} A mixed effects model may give a lower residual variance – like in this case – because information of incomplete data is recovered. R lmer() is set up according to the EMA’s ‘Method B’ which treats subject as a random effect but is not a true mixed effects model. Given only for completeness (for the code see below). Mixed effects in R are a swamp. The closest I could get was this:
Extracting the required stuff is beyond me. We had many discussions already in the forum and the verdict was “mixed effects models for BE are not possible in R”. So if you are happy with a fixed effect model (many jurisdictions like the EMA), fine. If you have to deal with the FDA or Health Canada, maybe not.
— Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
mittyri ★★ Russia, 20190324 00:13 (1807 d 02:32 ago) @ Helmut Posting: # 20073 Views: 17,980 

Dear Helmut, ❝ A request to the SASians of the forum: Please run Proc MIXED with DDFM=SATTERTHWAITE of the pooled data and both DDFMoptions on the data where S is excluded. Here's my (naive) attempt <...> Results:
Option Label Estimate StdErr DF tValue Probt Alpha Lower Upper pe lowerCL upperCL — Kind regards, Mittyri 
Helmut ★★★ Vienna, Austria, 20190324 00:36 (1807 d 02:09 ago) @ mittyri Posting: # 20074 Views: 17,802 

Hi mittyri, ❝ ❝ A request to the SASians of the forum: Please run Proc MIXED with DDFM=SATTERTHWAITE of the pooled data and both DDFMoptions on the data where S is excluded. ❝ Here's my (naive) attempt THX a lot. I added your result to the comparison above. — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
mittyri ★★ Russia, 20190324 00:50 (1807 d 01:55 ago) @ Helmut Posting: # 20076 Views: 17,592 

Dear Helmut, since you are very quick with answers even Saturday night, I'm not editing my previous post but commenting it here ❝ Extracting the required stuff is beyond me. We had many discussions already in the forum and the verdict was “mixed effects models for BE are not possible in R”. Hmmm... If I remember correctly we were struggling with FDA model where sophisticated 'repeated' statement exists. We successfully crosschecked EMA method B (simple mixed effects model with a Subject as random effect). So linear mixed effects models are possible until FDAstyle is required . — Kind regards, Mittyri 
Helmut ★★★ Vienna, Austria, 20190324 02:23 (1807 d 00:22 ago) @ mittyri Posting: # 20077 Views: 17,905 

Hi mittyri, ❝ If I remember correctly we were struggling with FDA model where sophisticated 'repeated' statement exists. ❝ We successfully crosschecked EMA method B (simple mixed effects model with a Subject as random effect). Are you reminding Detlew and me obout our TODOlist? ❝ So linear mixed effects models are possible until FDAstyle is required . Yes, but that’s the point. However, recycled our code: library(lmerTest) # (requires lme4, Matrix) Gives:
Similar ≠ same. Though the DFs are slightly different, the CIs look only identical due to rounding. — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
Brus ★ Spain, 20210609 18:48 (998 d 08:57 ago) @ Helmut Posting: # 22398 Views: 8,310 

Dear Helmut, I have a question on this topic that you mentioned a long time ago: ❝ A 6×3 design is needed in order to ‘extract’ two 2×2 tables, which are also balanced. Although the full 6×3 table will be used in the analysis of AUC and C_{max}, you will need these 2×2s for the nonparametric analysis of t_{max} (unfortunately there’s no confidence interval based nonparametric method available for more than two formulations/periods). The asterisks ❝
❝ ^ balanced ^ balanced ❝ A common mistake is to design the study as a set of 3×3 latin squares, which will lead (especially if the sample size is small and in the case of drop outs) to extremely imbalanced data sets: ❝
❝ ^ imbalanced ^ imbalanced According to what you said, 6 sequences had to be done with a 3way study to get balanced sequences on the two 2x2 tablets. But, with following approach, balanced sequences would also be achieved: ABC BAC CAB CBA It's possible? What problems would it have? It is not williams square design, right? Best regards, 
Helmut ★★★ Vienna, Austria, 20210609 19:48 (998 d 07:58 ago) @ Brus Posting: # 22399 Views: 8,217 

Hi Brus, ❝ I have a question on this topic that you mentioned a long time ago: […] Please read the entire thread. What I posted in 2007 is obsolete. See also this article. ❝ According to what you said, 6 sequences had to be done with a 3way study to get balanced sequences on the two 2x2 tablets. But, with following approach, balanced sequences would also be achieved: ❝ ❝ ABC ❝ BAC ❝ CAB ❝ CBA That’s not balanced for carryover. However, since we cannot correct for unequal carryover anyway (see there), we have to avoid it by design, i.e., sufficiently long washouts between treatment periods. ❝ It's possible? What problems would it have? What is your idea behind such a design? Actually a simple Latin Square (ABCBCACAB) would do as well. If you want to make European assessors happy, opt for the 6sequence Williams’ design. Most important: Evaluate the study by the ‘Two at a Time’ approach. ❝ It is not williams square design, right? Correct. — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
Brus ★ Spain, 20210610 16:33 (997 d 11:13 ago) @ Helmut Posting: # 22402 Views: 8,063 

Hi Helmut, ❝ That’s not balanced for carryover. However, since we cannot correct for unequal carryover anyway (see there), we have to avoid it by design, i.e., sufficiently long washouts between treatment periods. Why? IMHO, in my 3x4x6 design proposal, ABC BAC CAB CBA Eeach formulation is administered once per subject, occurs the same number of times in each period, and any formulation is preceded equally often by each of the other formulations. In this way it is a balanced design. What benefit does williams design bring? ❝ What is your idea behind such a design? Make a balanced design with as few sequences as possible. ❝ Actually a simple Latin Square (ABCBCACAB) would do as well. If you want to make European assessors happy, opt for the 6sequence Williams’ design. Will Simple Latin Square be accepted by EMA assessors? Will my proposal design be accepted by EMA assessors? Best regards, 
Helmut ★★★ Vienna, Austria, 20210610 17:03 (997 d 10:43 ago) @ Brus Posting: # 22403 Views: 8,437 

Hi Brus, ❝ ❝ That’s not balanced for carryover. ❝ ❝ Why? Sorry (see below). ❝ Eeach formulation is administered once per subject, occurs the same number of times in each period, and any formulation is preceded equally often by each of the other formulations. In this way it is a balanced design. So far, so good. But: In your design you have to assume equal period effects. Let’s count how many times the treatments are administered in each period.
❝ What benefit does williams design bring? It’s also balanced for period effects, which is really important.
❝ ❝ What is your idea behind such a design? ❝ ❝ Make a balanced design with as few sequences as possible. Got it. ❝ ❝ Actually a simple Latin Square (ABCBCACAB) would do as well. If you want to make European assessors happy, opt for the 6sequence Williams’ design. ❝ ❝ Will Simple Latin Square be accepted by EMA assessors? Possible. Statistically nothing speaks against it. We don’t have carryover in the model and it’s balanced for period effects. ❝ Will my proposal design be accepted by EMA assessors? Unlikely. If you have true period effects, your estimates will be biased and you have no means to correct it. — Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
vezz ☆ Erba (CO), Italy, 20210610 19:23 (997 d 08:23 ago) @ Helmut Posting: # 22404 Views: 8,166 

❝ Unlikely. If you have true period effects, your estimates will be biased and you have no means to correct it. This is not to support the design proposed by Brus, but it seems to me that with those sequences we should still be able to obtain unbiased estimates of treatment effect also in presence of a period effect (based on the model suggested by the EMA Guideline on BE). However, for some treatment comparisons the SE will be larger compared to a Latin square design with the same sample size. I look forward for your thoughts! — Kind regards, Stefano 
Helmut ★★★ Vienna, Austria, 20210610 22:14 (997 d 05:31 ago) @ vezz Posting: # 22405 Views: 8,086 

Hi vezz, ❝ ❝ Unlikely. If you have true period effects, your estimates will be biased and you have no means to correct it. ❝ ❝ This is not to support the design proposed by Brus, but it seems to me that with those sequences we should still be able to obtain unbiased estimates of treatment effect also in presence of a period effect (based on the model suggested by the EMA Guideline on BE). Nope. Let’s consider a simple example. A = B = C = 1, no carryover, additive model.
— Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
vezz ☆ Erba (CO), Italy, 20210611 11:11 (996 d 16:35 ago) @ Helmut Posting: # 22406 Views: 8,095 

Hi Helmut, in your example you are assuming a model with no period effect, while I am considering the usual model for the analysis of BE studies, including the period effect. With the period effect included in the model, the estimates of treatment effect are unbiased also using Brus' design. — Kind regards, Stefano 
Brus ★ Spain, 20210611 14:19 (996 d 13:27 ago) @ vezz Posting: # 22408 Views: 8,202 

Dear Vezz and Helmut, I think I understood Helmut. With my design (3x4x3), the carryover effect would be covered, since it generates balanced sequences for twototime comparisons, and any formulation is preceded equally often by each of the other formulations, but it does not cover the period effect, since that not all treatments are the same number of times in all periods (for example, in my design treatment "A" is administered 1 time in period 1, twice in period 2 and 1 time in period 3. In this way, in the event that there was a period effect, it would not be offset by the design and it would affect the results. Right? As Vezz says, if you consider period effect in your model and the software itself already incorporates that correction in case there is period effect, you could have unbiased results, in fact, considering the period effect and the rest of possible effects (sequence, treatment,...) in the model, any design would be acceptable, right ? But at this point, a question comes to mind, if our software is capable of calculating the period, sequence, group, and treatment effect, what need is there to make balanced designs? In this way IMHO, and if I'm not mistaken, after kindly discussing it, I think the ideal option for a 3 treatment crossover study is 3x6x3 williams design. Best regards, 
Relaxation ★ Germany, 20210611 14:42 (996 d 13:03 ago) @ vezz Posting: # 22409 Views: 8,000 

Hi Stefano. I was a little bit hesitant to answer, as I am and likely will remain a noob in the statistical background. Maybe I also misunderstand your question, which is quite possible. But there are a lot of people around here to correct the answer, if necessary, and its SOPFriday... I think I get your point, but I also think that the period effect is implemented above. The numbers already include both as mean=µ+π = treatment + period effect. As an example: the term s2p2 above is not the treatment mean for A in sequence 2 and period 2 (=1), it is the observed mean and includes the effect of period 2 (10.5=0.5). If we would like to split that up into more detail, when sxpy is the treatment only (defined by combination of sequence and period), we can add Py as the period effect for more clarity. Then: AC = ((s1p1 + P1) + (s2p2 + P2) + (s3p2 + P2) + (s4p3 + P3))  ((s1p3 + P3) + (s2p3 + P3) + (s3p1 + P1) + (s4p1 + P1)). See, as in Helmuts post, the period effects are implemented. But in the discussed design (P1+P2+P2+P3 = 0.5) and (P3+P3+P1+P1 = 1) do not completely eliminate each other as they would in a Latin Square. Thus treatment difference is biased (to +0.5 or 3.5/5=0.7) and not easily corrected. Now, lets both wait and see, whether I got it right . Best regards, Relaxation. 
Helmut ★★★ Vienna, Austria, 20210611 17:41 (996 d 10:04 ago) @ Relaxation Posting: # 22410 Views: 8,030 

Hi Relaxation and all, ❝ Now, lets both wait and see, whether I got it right . You did. In simple terms: In – any – crossover period effects mean out if (and only if) the number of treatments in each period are equal because then they are affected to the same extent. That’s one of the reasons why we design studies for balanced sequences. Due to dropouts sequences may be imbalanced, which biases the outcome. However,
— Diftor heh smusma 🖖🏼 Довге життя Україна! _{} Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes 
vezz ☆ Erba (CO), Italy, 20210611 18:56 (996 d 08:49 ago) @ Relaxation Posting: # 22411 Views: 8,060 

❝ I was a little bit hesitant to answer, as I am and likely will remain a noob in the statistical background. Maybe I also misunderstand your question, which is quite possible. ❝ But there are a lot of people around here to correct the answer, if necessary, and its SOPFriday... ❝ ❝ I think I get your point, but I also think that the period effect is implemented above. ❝ The numbers already include both as mean=µ+π = treatment + period effect. Hi Relaxation and Helmut, the period effect was taken into account by Helmut when generating the data, but not when analysing them. I am not able to explicitly describe each single step behind the estimation of the regression model (this would take some time!), but I will share with you the SAS code replicating the example. Some notes:  Treatments are coded as 123 instead of ABC.  Without loss of generality, I am assuming 6 subjects per sequence and with the PARMS statement in the MIXED procedure I keep the residual variance fixed at 10.  In the first MIXED procedure a model not including the period effect is estimated, while in the second one the model includes the period effect. data a01 (drop=i t1t3); Estimated treatment differences (SE) by the first model (no period effect):  A vs. B: 3.89E16 (0.9129)  A vs. C: 0.3750 (0.9129)  B vs. C: 0.3750 (0.9129) Estimated treatment differences (SE) by the second model (period effect included):  A vs. B: 3.7E16 (0.9129)  A vs. C: 753E17 (1.0206)  B vs. C: 79E16 (1.0206) We may notice that in the second model:  The estimate of the treatment effect is always practically 0, therefore unbiased.  SEs are not identical as the design is not balanced for period. — Kind regards, Stefano 