Bioanalytical Method (continued) / NSRB / PRC Narrative / Natural Hx / Population Analysis / Quantitative BM Criteria

[Sarah]. It does seem like once the custom probe synthesis is triggered, it is best to ship directly to TSHO, along with the positive and negative controls.

[Nishihara] If the team agree with your proposal and you can prepare the extra probe, before the monkey testing, TSHO DMPK can probably provide any ISH data whether mRNA is distributed in neurons of mouse SN using a in-house super-resolution microscope

[xin]

• Sarah introduced the RNAscope probe for our human condo optimized PRKN developed by ACDBio.
• 3 options for conducting probe validation study
  1. Conduct it in TBOS DSRE (Action: Yasuno/Saku to check with TBOS DSRE for their capability and timeline to do the study)
  2. Conduct it in ADDP (Action: Mitsuhiro/Kimio to check with ADDP for their capability and timeline in parallel)
  3. Conduct it in TSHO DMPK if option 1 and 2 are not feasible. TSHO DMPK don’t have the experience, will need more sample for pilot study.
• Samples: WT mouse and Monkey as negative control and AAV-treated PRKN KO mouse as positive control (Action: Takeshi to organize the samples for validation study)
• After we get ready for doing the validation study and also for the samples, Sarah will go ahead to order the probes and shipped to lab where we will perform the study. It takes 10 days from ordering to shipping.

Yasuno, Hironobu: TBOS DSRE has capability for RNAscope using the Leica Bond RX system according to the automated ISH protocol for RNAscope 2.5 LS Reagent Kit-Brown (ACDBio).

It depends on their availability, but generally, once they receive the probes and tissue samples, it takes about 2 weeks to prepare the stained slides.

Sarah: Did you want us to send from Takeda, or did you want ACDBio to ship directly to Shonan

Date / labels	Note
20230124 Yasuno (?)	The expression of antisense probe (CPG5-2) in PRKN KO mouse unilateral SN (injection site) is confirmed in PK041 treated group while not observed in vehicle treated group. In WT mouse, CPG5-2 expression is observed in unilateral SN (Injection site). In monkey SN, there is also no expression of CPG5-2. All these results suggest a good sensitivity and selectivity towards mouse/monkey PRKN.
	In the coming NHP study, RNA probe will be mainly used and DNA probe still needs to be validated and will be used in future studies.
20230227 Naohiro Narita, Kimio Iohyama, Kazuko Watanabe )	high specificity toward human parkin vs monkey parkin or mouse parkin. In rat brain: AAV9-PK041 (Low dose: 2e11 vg?) in SN → 17X higher hPRKN level

JSRB

[Parkin GT_NSRB for PRC PE_2021_1119.pptx]

• Working document of Parkin GT PRC narrative:
  • [Working document_PRC_Narrative_Parkin_PD_GT.docx]
  • draft to Ceri by Nov 12. (Before that we will send a draft version to PRC office by Nov 5).
  • [Working document_3rd draft_PRC narrative_Parkin GT.docx]
• [Anticipated questions in PRC.xlsx]

Our aim is to finalize 1st draft by Oct 11.

	Jaya	Jaewon	Animal model
presymptomatic or prodromal	disease progression modelling could be done with natural history data to identify variables that predict onset of symptoms, such as age of onset in affected family members or particular PRKN mutation variants. Identified presymptomatic biallelic PRKN-PD carriers could be monitored, perhaps through periodic DaT scans or passive digital monitoring, and then treated when they demonstrate prodromal symptoms or substantial loss of dopaminergic signal.
symptomatic biallelic PRKN-PD and demonstrate reversal of clinical symptoms	Na	The change of Datscan may be different from that of MDS-UPDRS. SO naturia Hx data worth.	6OHDA mice → AAV-Parkin → locomotion? 필요,
levodopa-induced dyskinesia

Post-PE

action item sheet .

[Parkin GT_Team Action Items for PRC e-submission.docx]

submission document

[Parkin GT_Supplement_e-submission_PRC narrative.docx]

Population analysis

MJFF GGPD study involves 89 centers worldwide in 42 countries. Full MJFF deck attached.

MJFF GGPD Project - Subjects

	PD+	PD-	How many can be recontacted?	Centers
Total	3185	703	3013 (75%)	89
Parkin (biallelic)	429	1	383 (89%)	60
Parkin (monoallelic)	217	89	202 (66%)	39
PINK1 (biallelic)	75	1	66 (87%)	23
PINK1 (monoallelic)	56	45	82 (81%)	13

→ 3,888 subjects

1. ROPAD study – 0.8% PRKN mutation carriers (biallelic and monoallelic) out of 13,000 PD subjects genotyped (any age of onset)

About 700 Parkin and PINK1 patients from 89 centers identified through the MJFF Parkin/PINK1 effort (biallelic and monoallelic), of whom 2/3 can be re-contacted.

Quantitave BM criteria:

		Dysregulation in patients	Needed change
기술		will be informed by animal study, natual Hx study	a quantitative relationship between the change of the biomarkers and treatment response will be modelled using a rat disease model in combination with 6-OHDA, MPTP treatment. (is there a linear correlation? What's the degree of correlation? Is there a threshold?)
CSF Parkin protein		To be verified (cf? brain ↓ 70% )	(GT : ↑ X1)
Ps65-ub		↓ 100% (to be verified)
MC1		? to be verified, (cf ↓ 30% MC1 activity),
Datscan	No need to mention

For clinical trials with ‘PD patients with PRKN mono-allelic mutation’ and ‘PD patients with low Parkin protein levels’, the same biomarkers that are currently in development for ‘PD patients with PRKN bi-allelic mutation’ will be used: transgene expression (CSF Parkin protein), pharmacodynamic effect (CSF ps65-ub) and disease progression (DaTSCAN).

20220119 기술본: To develop a biomarker that can be used for selecting PD patients with low Parkin protein levels, the team will establish a correlation of the level of biomarkers (eg. Parkin protein and ps65-ub) between the target brain regions (ie nigrostriatum), CSF and plasma using postmortem samples of PD patients. In order to define the biomarker quantitative targets, a quantitative relationship of the treatment-induced changes in the biomarkers and DA neurodegeneration will be modelled using a rat disease model. If a novel Parkin PET tracer is successfully developed, it can be used for the patient selection.

20220130 다음처럼 바뀌었네.

Uncertain Spans

location	transcription	uncertainty
Specificity row 2 date	20230227 Naohiro Narita, Kimio Iohyama, Kazuko Watanabe )	The trailing ) after Watanabe is visible in the source; the closing parenthesis has no matching open paren in the visible cell.
NSRB header	JSRB	A single-line label appears at the top of the NSRB section; the linked file is Parkin GT_NSRB for PRC PE_2021_1119.pptx, which suggests the heading may be an OCR artifact for NSRB, but the visible glyph reads as JSRB.