SEND Example.In vitro Micronucleus Assay - includes smoke regimen and device identifier

This is an example showing trial design and results data of Study #123 for the determination of the in vitro genotoxicity potential of 10 tobacco products in the in vitro Micronucleus Assay

red font - indicates potential for CT code lists
green font - links to other domains
purple font - to be discussed

For the purposes of team review of the example data, the report is included in this section:

Sample data #1: Determination of the in vitro genotoxicity potential of 10 tobacco products in the in vitro Micronucleus Assay

Study info: This study was performed to assess the in vitro genotoxicity of 10 different tobacco products containing 1% to 2% nicotine. The genotoxic potential was determined using the in vitro micronucleus test with TK6 lymphoblastoid suspension cells. The study was conducted in compliance with the following documents:

OECD TG 487 (2010): Guideline for the testing of chemicals: In vitro mammalian cell micronucleus test.
BL SOP 132: Determination of the in vitro genotoxicity of condensates from tobacco products and ingredients for tobacco products / electronic vapour products – in vitro micronucleus test (IVM) with TK6 cells.

The cells were exposed to increasing dose levels of tobacco product using short term treatment in the presence and absence of an external metabolic activation system (ST+/-S9 mix) as well as a long term treatment in the absence of an external metabolic activation system (LT-S9 mix).

Toxicity was calculated as relative increase in cell count (RICC), relative cell count (RCC) and relative population doubling (RPD). RPD is the cytotoxicity measure used for the assessment. RICC and RCC are also reported but not considered for the assessment.

Note: OECD GUIDELINE FOR THE TESTING OF CHEMICALS

Population Doubling = [log (Post-treatment cell number ÷ Initial cell number)] ÷ log 2

Conclusion: All tobacco product evaluated did not induce any signs of severe toxicity or genotoxicity in any of the treatments and do not fulfil the criteria to be classified as genotoxic.

Reported results: (only listed the result of one tobacco product as an example)

Statistical analysis

Adjusted p-values calculated for the micronucleus frequencies for every dose level as compared to the corresponding solvent control after ST in the presence and absence of S9 mix as well as LT in the absence of S9 mix. One way ANOVA with posthoc Dunnett’s test for multiple comparisons for the dose response and a two tailed unpaired student’s t-test for the comparison of positive and negative control were used. The difference between the samples is considered statistically significant at p≤0.05.

Treatment	Test date	Concentration [µg/ml]	Fold increase of MN over background	Adjusted p- value	Significant increase Y/N
ST+ S9	25.05.2022	1250	0.9	0.9973	N
		2500	0.9	0.9517	N
		3750	0.6	0.2654	N
		5000	0.9	0.9895	N
		CPA [5µg/ml]	3.1	0.0003	Y
ST-S9	25.05.2022	1250	1.6	0.6408	N
		2500	1.3	0.8995	N
		3750	2.9	0.0106	N
		5000	1.0	> 0.9999	N
		Bleo [0.2µg/ml]	3.8	0.0002	Y
LT-S9	25.05.2022	1250	0.7	0.5958	N
		2500	1.3	0.6378	N
		3750	0.5	0.1146	N
		5000	1.1	0.9929	N
		Bleo [0.5µg/ml]	2.9	0.0047	Y

Bleo: Bleomycin; CPA: Cyclophosphamid A; statistically significant increases are highlighted in bold.

Assumption: The intent of this dataset is to provide a summary of trial (study) information. This is not subject-level data.
Assumption: A Trial (study) can have more than one assay type
Assumption: ASSAYID value of ALL indicates that it applies to all assays in the study
Assumption: SPDEVID and DUREFID should only be in TX (with the same value for all sets if there is only one device used)
Assumption: SPDEVID (sponsor defined device identifier) should be added at the trial set level (tx.xpt) - we can discuss if this should be in TS when there is only one device for a study
- This allows for studies where there are multiple products and different product(s) per trial set, one record for each product that is being tested in each particular trial set (tx.xpt)
Do we need to document anything about extraction process steps from the smoking machine to a test material for an assay (MNvit) (ie, rinsing a filter??)
Concentration = 0 is negative control, Bleo and CPA are positive controls - confirm

Row	STUDYID	ASSAYID	DOMAIN	TSSEQ	TSGRPID	TSPARMCD	TSPARM	TSVAL	TSVALNF
1	123	MNvit	TS	1		GLPTYP	Good Laboratory Practice Type	FDA
2	123	MNvit	TS	2		GLPTYP	Good Laboratory Practice Type	OECD
3	123	MNvit	TS	1		STSTDTC	Study Start Date	2022-05-25
4	123	MNvit	TS	1		STITLE	Study Title	Determination of the in vitro genotoxicity potential of 10 tobacco products in the in vitro Micronucleus Assay
5	123	MNvit	TS	1		SNDIGVER	SEND Implementation Guide Version	TOBACCO IMPLEMENTATION GUIDE VERSION 1.0
6	123	MNvit	TS	1		SNDCTVER	SEND Controlled Terminology Version	SEND Terminology 2021-09-30
7	123	MNvit	TS	1		SSPONSOR	Sponsor Organization	Example Sponsor Inc.
8	123	MNvit	TS	1		SPREFID	Sponsor's Study Reference ID		NOT APPLICABLE
9	123	MNvit	TS	1	1	TSTFNAM	Test Facility Name	Example Tox Lab Name
10	123	MNvit	TS	1	1	TSTFLOC	Test Facility Location	10 Somewhere Street, Montgomery, AL 10000
11	123	MNvit	TS	1	1	TFCNTRY	Test Facility Country	USA
12	123	MNvit	TS	1	1	STDIR	Study Director	Dr. R. Smith
13	123	MNvit	TS	1		GLPFL	GLP Flag	Y
14	123	MNvit	TS	1		ASTD	Assay Standard	OECD Test No. 487
15	123	MNvit	TS	1		ASTDV	Assay Standard Version	2016-07-29
16	123	MNvit	TS	1		SSTYP	Study Type	GENOTOXICITY IN VITRO
17	123	MNvit	TS	1		SSSTYP	Study Sub Type	In Vitro Micronucleus
18	123	MNvit	TS	1		SPECIES	Species	Homo Sapiens
19	123	MNvit	TS	1		TESTSYS	Test System	TK6 Lymphoblastoid Suspension Cells (also need Rat cells as an option) In OECD 487: Cultured primary human peripheral blood lymphocytes (5)(19)(42)(43) and a number of rodent cell lines such as CHO, V79, CHL/IU, and L5178Y cells may be used (18)(19)(20)(21)(22)(25)(26)(27)(28)(30).

(This is a copy from the SDTM Example. Cigarette Design Parameters, will need scription team)

Rows 1-4:	Show the records for the product identifiers for the tobacco product identified in SPTOBID. These records are categorized as product identifiers by TOCAT = PRODUCT IDENTIFIERS.
Rows 5-10:	Show the records for the product descriptors for the tobacco product identified in SPTOBID. These records are categorized as product identifiers by TOCAT = PRODUCT DESCRIPTOR.

to.xpt

Row	STUDYID	DOMAIN	SPTOBID	TOSEQ	TOTESTCD	TOTEST	TOCAT	TOORRES	TOORRESU	TOSTRESC	TOSTRESN	TOSTRESU
1	TOB07	TO	CIG01a	1	TBPRDCAT	Tobacco Product Category	PRODUCT IDENTIFIER	Cigarette		Cigarette
2	TOB07	TO	CIG01a	2	TBPRSCAT	Tobacco Product Subcategory	PRODUCT IDENTIFIER	Filtered, Combusted		Filtered, Combusted
3	TOB07	TO	CIG01a	3	MANUF	Manufacturer	PRODUCT IDENTIFIER	Joes Cigs USA		Joes Cigs USA
4	TOB07	TO	CIG01a	4	TRADENAM	Trade Name	PRODUCT IDENTIFIER	Treetop Menthol King Size		Treetop Menthol King Size
5	TOB07	TO	CIG01a	5	PACKTYP	Package Type	PRODUCT DESCRIPTOR	HARD PACK		HARD PACK
6	TOB07	TO	CIG01a	6	PRDQUAN	Product Quantity	PRODUCT DESCRIPTOR	20	CIGARETTE	20	20	CIGARETTE
7	TOB07	TO	CIG01a	7	LENGTH	Length	PRODUCT DESCRIPTOR	86.0	mm	86.0	86.0	mm
8	TOB07	TO	CIG01a	8	CIRCUMF	Circumference	PRODUCT DESCRIPTOR	26.0	mm	26.0	26.0	mm
9	TOB07	TO	CIG01a	9	VENTLTN	Ventilation	PRODUCT DESCRIPTOR	10.0	%	10.0	10.0	%
10	TOB07	TO	CIG01a	10	CHARFLAV	Characterizing Flavor	PRODUCT DESCRIPTOR	MENTHOL		MENTHOL

During CT definition/reviews will decide appropriate TXPARM and TXVAL; Treatment duration may be controlled; For now, we just include good example values based on our experience
Assumption: The Trial Sets (TX) domain provides the list of distinct sets of subjects having different experimental factors, treatment factors, inherent characteristics, or distinct sponsor designations as specified in the trial design.
Where is TK6 cell type? is this test system (see below)
- needs to be allowed to vary down to the well level / result level
Do we need both SPDEVID AND DUREFID?
What are good values (realistic) for "SET"?
Check Essential Data list

A1:

A2:

Row	STUDYID	ASSAYID	DOMAIN	SETCD	SET (what sponsor calls it, more meaningful label for Tables)	TXSEQ	TXPARMCD	TXPARM	TXVAL
1	123	MNvit	TX	A1 (table 1, row 1, ST exposure with S9)	ST+S9C0	1	METACT	Metabolic Activation (this is the type of activation used)	+S9
2	123	MNvit	TX	A1	ST+S9C0	2	METACTFL	Y/N presence of metabolic activation (this indicates that metabolic activation was used)	Y
3	123	MNvit	TX	A1	ST+S9C0	3	TRTDMIN	Treatment Duration Minimum	3
4	123	MNvit	TX	A1	ST+S9C0	4	TRTDTRG	Treatment Duration Target	3.5
5	123	MNvit	TX	A1	ST+S9C0	5	TRTDMAX	Treatment Duration Maximum	4
6	123	MNvit	TX	A1	ST+S9C0	6	TRTDU	Treatment Duration Unit	HOURS
7	123	MNvit	TX	A1	ST+S9C0	7	RCVDMIN	Recovery Duration Minimum	23.5
8	123	MNvit	TX	A1	ST+S9C0	8	RCVDTRG	Recovery Duration Target	24
9	123	MNvit	TX	A1	ST+S9C0	9	RCVDMAX	Recovery Duration Maximum	24.5
10	123	MNvit	TX	A1	ST+S9C0	10	RCVDU	Recovery Duration Unit	HOURS
11	123	MNvit	TX	A1	ST+S9C0	11	INCBTMP	Incubation Temperature	37
12	123	MNvit	TX	A1	ST+S9C0	12	INCBTMPU	Incubation Temperature Unit	C
13	~~123~~	~~MNvit~~	TX	A1	~~ST+S9C0~~	13	~~MEDIA~~	~~Composition of Media~~	~~Sartorius HEK293~~
14	123	MNvit	TX	A1	ST+S9C0	14	HUMID	Atmospheric Relative Humidity Percent	50
15	123	MNvit	TX	A1	ST+S9C0	15	ATMCO2	Atmospheric CO2 Percent	5
16	123	MNvit	TX	A1	ST+S9C0		SPTOBID	Sponsor defined tobacco identifier	CIG01a
17	123	MNvit	TX	A1	ST+S9C0		EXPTYP	Exposure Type (See TIG NC workstream minutes 30-Jan here: Nonclinical)	Submerged
	123	MNvit	TX	A1	ST+S9C0		SAMTYP	Sample Type (e.g. TPM, GVP, whole aerosol, whole smoke conditioned media, aqueous extracts, etc., see notes here: Nonclinical)	Total Particulate Matter in DMSO
18	123	MNvit	TX	A1	ST+S9C0	6	INTRVN	Name of the Intervention Article can be: Tobacco ProdA, Bleomycin (positive control) or Cyclophosphamid A (positive control)	Tobacco ProdA
19	123	MNvit	TX	A1	ST+S9C0	7	ITVTYPE	type of intervention article choices of values: product; negative control; positive control	Negative Control
20	123	MNvit	TX	A1	ST+S9C0	8	ITVCONC	Concentration of intervention article	0
21	123	MNvit	TX	A1	ST+S9C0	9	ITVCONCU	Concentration Unit	ug/ml
22	123	MNvit	TX	A1	ST+S9C0	10	SPDEVID	Sponsor defined device identifier	PUFFMASTER3K
23	123	MNvit	TX	A1	ST+S9C0	11	DUREFID	Smoke Regimen	Medium Intensity Regimen
24	~~123~~	~~MNvit~~	TX	A1	~~ST+S9C0~~	11	~~SMKEXSYS~~	~~Smoke Exposure System~~	~~Cambridge filter pad, eluted in DMSO~~
25	123	MNvit	TX	A2 (table 1, row 2, ST exposure with S9 at concentration 1250)	ST+S9-1250	1	METACT	Metabolic Activation (should there be two parms? Presence, type)?	+S9
26	123	MNvit	TX	A2	ST+S9-1250	2	METACTFL	Y/N presence of metabolic activation (this indicates that metabolic activation was used)	Y
27	123	MNvit	TX	A2	ST+S9-1250	3	TRTDRTRG	Treatment Duration target. (how do we show 3-6 hour range? start/end, target and tolerance?, one text field not-analyzable)	3
28	123	MNvit	TX	A2	ST+S9-1250	4	TRTDRTOL	Treatment Duration Tolerance
29	123	MNvit	TX	A2	ST+S9-1250	5	TRTDURU	Treatment Duration Unit (this is for both TRTDURT, TRTDURTOL)	H
30	123	MNvit	TX	A2	ST+S9-1250	6	INTRVN	name of the intervention article	Tobacco ProdA
31	123	MNvit	TX	A2	ST+S9-1250	7	ITVTYPE	type of intervention article	Product
32	123	MNvit	TX	A2	ST+S9-1250	8	ITVCONC	Concentration of i a	1250
33	123	MNvit	TX	A2	ST+S9-1250	9	ITVCONCU	Concentration Unit	ug/ml
34	123	MNvit	TX	A2	ST+S9-1250	10	SPDEVID	Sponsor defined device identifier	PUFFMASTER2023
35	123	MNvit	TX	A2	ST+S9-1250	11	DUREFID	Smoke Regimen	High Intensity Regimen
...

We use DU for smoking regimen. Note that a separate DI dataset will be needed to show identifying parameters of the "PUFFMASTER3K" smoking machine

Details of the smoking regimen are represented as device in-use properties, linked to the stability data in PT above by matching values of PTREFID/DUREFID = "Medium Intensity Regimen" (We will update with a realistic value for the regimen, with input).
Smoking regimen is represented in --REFID (we made up a value of "Medium Intensity Regimen"; we can update with something realistic)
The smoking regimen is carried out by the smoking machine/device shown in SPDEVID, Sponsor defined device identifier, "PUFFMASTER3K"

du.xpt

Row	STUDYID	DOMAIN	SPDEVID	DUSEQ	DUREFID	DUGRPID	DUTESTCD	DUTEST	DUORRES	DUORRESU	DUSTRESC	DUSTRESN	DUSTRESU
1	123	DU	PUFFMASTER3K	1	Medium Intensity Regimen		PUFFPROF	Puff Profile	SQUARE		SQUARE
2	123	DU	PUFFMASTER3K	2	Medium Intensity Regimen		PUFFDUR	Puff Duration	1.25	sec	1.25	1.25	sec
3	123	DU	PUFFMASTER3K	3	Medium Intensity Regimen		PUFFINT	Puff Interval	3	PUFF/min	3	3	PUFF/min
4	123	DU	PUFFMASTER3K	4	Medium Intensity Regimen		PUFFBLCK	Puff Block	25	%	25	25	%
5	123	DU	PUFFMASTER3K	5	Medium Intensity Regimen		NUMPUFF	Total Number of Puffs	200	PUFF	200	200	PUFF
6	123	DU	PUFFMASTER3K	6	Medium Intensity Regimen		PUFFVOL	Puff Volume	10	mL	10	10	mL
7	123	DU	PUFFMASTER3K	7	Medium Intensity Regimen		PUFFRNG	Puff Range	100-200		100-200
8	123	DU	PUFFMASTER3K	8	Medium Intensity Regimen	1	PUFFPAUS	Puff Pause	60	s	60	60	s
9	123	DU	PUFFMASTER3K	9	Medium Intensity Regimen	1	PUFFPINT	Puff Pause Interval	10	PUFF	10	10	PUFF
10	123	DU	PUFFMASTER2023	1	Canadian Intense Regime		PUFFPROF	Puff Profile	SQUARE		SQUARE
11	123	DU	PUFFMASTER2023	2	Canadian Intense Regime		PUFFDUR	Puff Duration	2.00	sec	2.00	2.00	sec
12	123	DU	PUFFMASTER2023	3	Canadian Intense Regime		PUFFINT	Puff Interval	4	PUFF/min	4	4	PUFF/min
13	123	DU	PUFFMASTER2023	4	Canadian Intense Regime		PUFFBLCK	Puff Block	0	%	0	0	%
14	123	DU	PUFFMASTER2023	5	Canadian Intense Regime		NUMPUFF	Total Number of Puffs	200	PUFF	200	200	PUFF
15	123	DU	PUFFMASTER2023	6	Canadian Intense Regime		PUFFVOL	Puff Volume	10	mL	10	10	mL
16	123	DU	PUFFMASTER2023	7	Canadian Intense Regime		PUFFRNG	Puff Range	100-200		100-200
17	123	DU	PUFFMASTER2023	8	Canadian Intense Regime	1	PUFFPAUS	Puff Pause	60	s	60	60	s
18	123	DU	PUFFMASTER2023	9	Canadian Intense Regime	1	PUFFPINT	Puff Pause Interval	10	PUFF	10	10	PUFF

di.xpt (copied v1.0 of medical devices IG)

This example is a copy of Example 1 from di.xpt in SDTMIG-MD v1.0 but with values for SPDEVID and DIVAL revised slightly.
Should I remove the FDA UDI (row 5) unless CTP has or plans to establish UDI values?

Example 1
This shows records for ~~two~~ three devices where the sponsor felt that the type, manufacturer, model number, and serial number were necessary for unique identification. In addition, there was a post-marketing UDI identifier available for the first device.

Rows 1-5 show the records for a device given a SPDEVID of PUFFMASTER3K
Rows 5-8 show the records for a device given a SPDEVID of PUFFMASTER2023
Rows 9-12 show

Row	STUDYID	DOMAIN	SPDEVID	DISEQ	DIPARMCD	DIPARM	DIVAL
1	123	DI	PUFFMASTER3K	1	DEVTYPE	Device Type	ENDS
2	123	DI	PUFFMASTER3K	2	MANUF	Manufacturer	Acme Machines
3	123	DI	PUFFMASTER3K	3	MODEL	Model Number	45-JFI
4	123	DI	PUFFMASTER3K	4	SERIAL	Serial Number	456789132-AXQ
5	123	DI	PUFFMASTER3K	5	FDAUDI	FDA Unique Device Identifier	456789123xyz
6	123	DI	PUFFMASTER2023	1	DEVTYPE	Device Type	SmokeMachine
7	123	DI	PUFFMASTER2023	2	MANUF	Manufacturer	Acme Machines
8	123	DI	PUFFMASTER2023	3	MODEL	Model Number	62-PLC
9	123	DI	PUFFMASTER2023	4	SERIAL	Serial Number	215964564-NFS
10	123	DI	UsualCTCigarette	1	DEVTYPE	Device Type	Combustible Tobacco Cigarette
11	123	DI	UsualCTCigarette	2	MANUF	Manufacturer	Philip Morris International
12	123	DI	UsualCTCigarette	3	MODEL	Model Number	Marlboro Red
13	123	DI	UsualCTCigarette	4	SERIAL	Serial Number	123456789

Entity IDs are unique within the assay (and a study can have many assays)
Replicates vs. Run
- need planned (ei.xpt) separate from the actual (gt.xpt)
- If tests run at the same time, they are replicates and are assigned the same RUNID
- If tests are run at separate times, they are runs and the RUNIDs are distinct
Different runs would be expected to have different ENID and GTDTC values. In ei.xpt, these are distinguished by different RUNID values.
Entities that are replicates of each other would be expected to
- have different ENID values
- be in the same trial set (SETCD values)
- have the same RUNID values

STUDYID	ASSAYID	DOMAIN	ENID (Entity ID)	SETCD (from TX)	RUNID	REPNUM (perm)	PLATEID (Plate ID)	COLID (Column number)	ROWID (Row number)
123	MNvit	EI	1-8-3	A1	1		1	8	3
123	MNvit	EI	2-8-3	A1	2
123	MNvit	EI	3-7-1	A2	1	1
123	MNvit	EI	2-7-1	A2	1	2

A1:

A2:

Different runs would be expected to have different ENID and GTDTC values. In ei.xpt, these are distinguished by different RUNID values.

Row	STUDYID	ASSAYID	DOMAIN	ENID	GTSEQ	GTTESTCD	GTTEST	GTCELLEV (cells evaluated)	GTORRES	GTORRESU	GTSTRESC	GTSTRESN	GTSTRESU	GTDTC
1	123	MNvit	GT	1-8-3	1	RICC	Relative Increase in Cell Count	154	0	%	0	0	%	2022-05-25
2	123	MNvit	GT	1-8-3	2	RCC	Relative Cell Count	154	0	%	0	0	%	2022-05-25
3	123	MNvit	GT	1-8-3	3	RPD	Relative Population Doubling	154	0	%	0	0	%	2022-05-25
4	123	MNvit	GT	1-8-3	4	MNCELLS	Micronucleated Cells	2205	15	Cells	15	15	Cells	2022-05-25
5	123	MNvit	GT	1-8-3	5	MNCELLS	Micronucleated Cells	2474	13	Cells	13	13	Cells	2022-05-25
6	123	MNvit	GT	1-8-3	6	MNCELLS	Micronucleated Cells	2758	17	Cells	17	17	Cells	2022-05-25
7	123	MNvit	GT	1-8-3	7	MNCELLS	Micronucleated Cells	2669	12	Cells	12	12	Cells	2022-05-25
8	123	MNvit	GT	1-8-3	8	AVGREL	Average Relative MN Frequency		0.57	%	0.57	0.57	%	2022-05-25
9	123	MNvit	GT		1	RICC	Relative Increase in Cell Count	134	15.7	%	15.7	15.7	%	2022-05-25
10	123	MNvit	GT		2	RCC	Relative Cell Count	134	13.0	%	13.0	13.0	%	2022-05-25
11	123	MNvit	GT		3	RPD	Relative Population Doubling	134	7.9	%	7.9	7.9	%	2022-05-25
12	123	MNvit	GT		4	MNCELLS	Micronucleated Cells	3266	20	Cells	20	20	Cells	2022-05-25
13	123	MNvit	GT		5	MNCELLS	Micronucleated Cells	2190	17	Cells	17	17	Cells	2022-05-25
14	123	MNvit	GT		6	MNCELLS	Micronucleated Cells	2758	13	Cells	13	13	Cells	2022-05-25
15	123	MNvit	GT		7	MNCELLS	Micronucleated Cells	2714	21	Cells	21	21	Cells	2022-05-25
16	123	MNvit	GT		8	AVGREL	Average Relative MN Frequency		0.66	%	0.66	0.66	%	2022-05-25
...

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SEND Example.In vitro Micronucleus Assay - includes smoke regimen and device identifier

SET (what sponsor calls it, more meaningful label for Tables)