| Possible reason | |
|---|---|
| Low SB on 6OHDA in both ARG and homogenate |
Low NLRP3 expression? (but well-detected in NLRP3 GoF homogenate) NLRP3 instability - eg. freeze/thaw NLRP3 instability - homogenization |
| Low NSB on 6OHDA in ARG | Toxin-induced morphology change |
Summary
| Bmax | Invicro | Date | Study type | Sample | Tracer | Competitor | Stimulation | Results | Next study | |||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| 202104 | PET SC 발표 | |||||||||||||||||||||||
| 202104 ARG | PET SC 발표 | 6OHDA rat (40 ug), striatum | 3H]MCC 950 | (~5 cc... ) | mouse and rat NLRP3 insect cell lysates | High (NLRP3) probe injected unilaterally to one of LPS-induced rat brain hemisphere across each of the analysed regions; 의외 NSB (i.e SB이 거의 없음) | Yamatake-san @TBOSS-Imaging will set up homogenate study to test out-of-house probes basic problems in vivo, vs vehicle, vs LPS NLRP3 GoF mice | |||||||||||||||||
| 1st | Optimization ARG | 6OHDA rat (n=1) | [3H]candidate 1 ([3H]TR06616126) | 10 / 100 nM | High (~10X) SB was observed in both the 1 site and 4 site 6 OHDA lesioned rat brains. There was a slight increase in SB signal on the lesioned hemisphere in relation to the sham lesioned hemisphere across each of the analysed regions | |||||||||||||||||||
| 2nd | Optimization ARG | the same dose of 6 OHDA spread across 4 sites | 3 H]MCC 950 | 6 nM | necrotic pockets, Binding was notably lower around the 6 OHDA lesion site in 'lesioned hemisphere across each of the analysed regions' | |||||||||||||||||||
| 20220118 | both Sec on 1 site and 4 site (40 ug) | 3 HPBR 28 | 10 nM | SB window was poor (<3X SB) in all regions. Little to no differences in SB observed between the sham and 6 OHDA lesioned hemispheres. Increased SB by ~5X with low doses (~50nM) at the 4 binding distribution of the binding resembled that seen with | ||||||||||||||||||||
| 202203 | Ad hoc 미팅 | 1st site (n=1?) | 3 HJPBR 28 | 1µL/4 site (rat) | various, Ashley CW | lesioned side / un-lesioned side: Very low NSB & Total at 100nM un-treated rat 70 ↑↑ p+ & Aconita, Ibotia c't qua & comita 시간 따라 binding range very low SB1, ?? / bonota ?? quat 2J쯤 ㅁ가 | ||||||||||||||||||
| Ad hoc 미팅 autopsy 인 human SC section | 3 H]MCC 950 | 2cc 30 ng / various PBR / 10 ug, 30 ug | MCC 950 | total 6m AD vs HCL n/sb / 92% reduction of [3H]PBR28 / 6m AD vs HC, n=10 each / [3H]PBR28 total signal in AD frozen 4-5x higher than that observed using 3 H]MCC 950 / HC brain: very low SB, ND brain: 60% and 75% reduction observed at 30nM / Binding of increasing concentrations of 3HMCC950 / SB window ranges from 20% to 55% with greatest displacement at 100nM (sub-nM 'future' 후 concentrations to reduce NSB) | ||||||||||||||||||||
| Postmortem PD samples |
i) Using the PD brain samples from NBB, the ARG study will be initiated. It will take ~2 months for the first Feasibility study including optimization and the results with NBB PD brain will be generated by the end of Apr. Tritiated MCC950 and TR126 will be used. ii) Further PD brain tissue from UK biobank will be obtained by the seasoned PD brain. The feasibility of PD brain will be evaluated in Jun. | |||||||||||||||||||||||
| Paul Rolzin |
Homogenate: confirming Invicro's study, with 6OHDA RATs homogenates. Bmax, J our codern, MCC950 Joshua Bliesath for homogenate
| |||||||||||||||||||||||
Triaging compounds
| Team: Anne Kanta (DMPK)-6OHDA rodent |
Hi Yamatake-san, Sorry for my delayed response. I had a shot chat with Masato and Mitsuhiro, and came up with some experimental condition to be conducted as a very first attempt. I will be summarizing that within this week and will discuss it with Jaewon first. In brief, we propose to focus on the rat sub-chronic LPS systemic injection model (4 times with 0.4 mg/kg i.p. as rereported) and my add another arm with increase of LPS dose. So the design would be
The dosing and sampling can be done by the Rabbics colleagues and PK evaluation will be organized by Mitsuhiro. • One missing capability in TSHO is ?? blot. So hope TSD team can support of NLRP3 protein expression change by the western blot. The challenges of the assay is that the NLRP3 protein appeared to be unstable in the extraction with lyse buffer. Wei has the proper experimental conditions for the WB. Now immediate next actions are 1) to confirm if the experimental protocol is covered by the currently approved one (Yoshiro, Masato), 2) determine the dose of probe compound started i.v. by referring to the LLOQ (Mitsuhiro), 3) confirm if TSD can support the WB experiment (Marianthi?), 4) Where NSTM discussion to finalize the protocol (Masato, Yoshiro, Jaewon) |
Dosing regimen
| Time point | WT rat | LPS (0.5 mg/kg) rat | LPS (2 mg/kg) rat | |
|---|---|---|---|---|
| 0.3 mg | 1 h | 3 | 3 | 3 |
| 2 h | 3 | 3 | 3 | |
| 3 mg | 1 h | 3 | 3 | 3 |
| 2 h | 3 | 3 | 3 |
Habituation/dosing schedule cartoon (SD rat 7-8 week, Day1 Habituation, Days 2-5 LPS (i.p.) arrows, Day 5 TR06647850 (i.v.) and Sampling marker — kept as evidence in body_r05_c01.jpg / body_r05_c02.jpg).
Footer: Private and confidential. For internal use only.
- readouts planned: i) brain penetration (Kp), ii) Bmax difference (PD vs WT), iii) regional distribution (SN, striatum?) of the probe (850) by LCMS? lii) block 한다고 안한다고
왼쪽에 대하여:
- Purpose?
- TO: (drug candidate 안 쓰니) 못 볼 것,
- SB?: i) KO 쓰면 NLRP3 protein-specific SB 볼 수 있을 텐데, 이걸 하진 않는 거고 ii) self-block? 그럼 competing agent (TR06647850)의 conc?
Next study: Triaging, Multiple tracer?, Competition assay (blocking)
TSHO PET-G (Yamatake) In vivo blocking study: 6-OHDA rat model @TCAL
Led by TSHO DMPK-G or TCAL DMPK
Overall control: NSTM (Yoshirou-san)
Candidate compounds: 1st: TR06673219 (TCAL series 3), 2nd: TR06647850 (Novartis Pat.)
Precursor: Synthesis by TSHO PET chemistry-G
Imaging (cold) / hot timeline:
- 202303: synthesis of precursor of 18F]TR06762852 ongoing
- 202304: ship [18F]TR06762852 precursor (5-10mg) and standard (25mg) to Tbos
- 202305: Paul Rolzin to surgery (4-level 6OHDA)
- 202306: Paul Rolzin ship 3-4 rats to Tbos
- 202306: label at Dana Farber → PET Study (baseline and blocking?)
Uncertain Spans
| location | transcription | uncertainty |
|---|---|---|
Summary tracer table / column placement | the wide multi-column tracer-experiment table has 9-10 columns including Bmax / Invicro / date / study type / sample / Tracer / Competitor / Stimulation / Results / Next study; per-row column placement is partly inferred from cell width visible in body_r01_c01.jpg. | low confidence on column-by-row placement. |
Summary tracer table / 202203 / Ad hoc 미팅 row | the result cell reads lesioned side / un-lesioned side: Very low NSB & Total at 100nM un-treated rat 70 ↑↑ p+ & Aconita, Ibotia c't qua & comita 시간 따라 binding range very low SB1, ?? / bonota ?? quat 2J쯤 ㅁ가; multiple Korean tokens are partly clipped. | low confidence on Aconita, Ibotia c't qua & comita token cluster. |
Triaging compounds / Now immediate next actions | the action items reference (Marianthi?) for WB experiment confirmation; preserved verbatim. | low confidence on whether Marianthi is the assignee or a question. |
Dosing regimen / TR06647850 row label | reads TR06647850 with strikethrough/cell-clip variation; preserved verbatim. | source typography preserved. |
Body bottom / Imaging timeline | reads 202303, 202304, 202305, 202306, 202306 (last entry duplicated); preserved verbatim. | low confidence on whether one of the 202306 entries should be 202307. |