Blog

Hello HOPEians,

Please find below the action items from the RF2 meeting on October 27, 2022. If you have any questions about the meeting or action items please reach out to Dr. Nadia Roan and Dr. Lish Ndhlovu. 

Updates:

Clinical Specimens

Lish gave an update on 26 patient samples from individuals on spironolactone.  Patients on eplerenone in the U.S. Military cohort were requested as a control; however, no PBMC was available.  50 controls were on amiloride.

Trevor is taking the lead for spironolactone clinical specimens.  Trevor Crowell and Brian will prepare the shipment inclusive of all controls. Collaboration between Denise/Lish/Susana -spironolactone PK in NHP samples (Matt in Thailand will send out the experimental plan to the working group).

Brazil cohort– Ana Carolina Soares mentioned that the study protocol still needs approval.  Spironolactone is not the only medication used, so enrollment is still ongoing.  No one is enrolled to date.

In vitro Experiments

Bulk RNA-seq on spironolactone-treated cells from Susana’s group may be useful in Nadia’s ex vivo primary single-cell RNA-seq study.  [~6-hour pre-treatment of spironolactone prior to infection].  Nadia Roan will continue to test PBMCs and tonsil samples. They are testing different concentrations of SP  and will block infection rates by 50% (not complete inhibition in order to leave some infected cells in spironolactone-treated condition). 

Ulrike gave an update on cell line models of HIV latency.  Generation of a cell line with a proviral reporter integrated into a region that is more heterochromatic (as reported in Elite Controllers).  CRISPR-based approaches will be used to generate these.  Alternatively, random infection and selection of clones where HIV integrates into heterochromatic regions.  Immortalization of primary cells with an hTERT plasmid – troubleshooting is underway.  Using primary T cell clones as a potential model, this needs to be optimized.

Luisa Mori is collecting data from patients treated with dCA ex vivo.  Susana discussed her group’s work on their dCA and spironolactone in combination experiments; J-Lat 10.6 cells.  The combination works better than each drug alone.  Different LRAs have been added to these conditions (8-hour LRA stimulation); GFP expression measured by flow cytometry was used as a readout.  Additionally, HIV mRNA (TAR, Gag-Pol, and Tat-Rev mRNA).  The results before Day 108 should be taken with caution due to technical issues. Focus should be placed on results past Day 108.

• Freeze/thaw greatly affects spironolactone efficacy and should be prevented by preparing more aliquots before initial freezing.  Breakdown products of Spironolactone do not work in HIV inhibition.

• Alina for Lish’s lab mentions that experimental time affects SP activity.  Susana’s experiment was 150 days. 

Susana also discussed Tat degrader TTX-881 plus Spironolactone combination experiments.  (all Tat degraders tested to date bind to Y26 residue on Tat).  After 12 days in culture, the combination condition inhibited HIV more than individually.  At 24 days, HIV was not as suppressed in this condition.  

Sonia Jablonski’s ongoing experiments:

1. Assessment of the impact of the tat degraders on tat degradation upon PROTAC cerblon knockdown in HEK293T cells

2. Optimization of the following pull downs: Tat ubiquitination, tat interaction with the components of the E3 ligase complex

3. Validate that the 3 tat degraders do not affect HIV integration

4. Ongoing analog synthesis of TT-22856 (Bannister’s lab)

5. TT-22856 was sent for pharmacokinetic analysis

6. Michael Corely / Alina Pang evaluate the 3 degraders in live cell imaging in tat 5A8 cells

Alina Pang shared some data from her dose-dependent inhibition of HIV reactivation after pre-incubation with 44863 experiments. The Tat degraders with preincubation for 5 days. She will test frozen samples for long-term effects. 

Actions:

• Denise – Send nonhuman primate studies/ PK experiment plan to the group soon

• Ana Leda – give DNA to Lish to look at the methylation state. 

Announcements:

• If you are attending the Miami Meeting and would like to join the HOPE dinner on 12/12 Please let Robert and me know. 

• The HOPE Collaboratory-wide meeting is on November 21st.

• The next RF2 Meeting is December 7th. Please come prepared with some updates. 

• Please Acknowledge HOPE & the NIH in any publications or presentations and notify the Program Manager. Here are some examples of what you can include. 

“Research reported in this publication was supported by the NIAID of the National Institutes of Health under award number UM1AI164559, with co-funding support from NIDA, NIMH, NHLBI, the NIDDK, and the NINDS. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.”

“This research was supported by NIAID award number UM1AI164559, co-funded by NHLBI, NIDA, NIMH, NINDS, and NIDDK.”

Hello HOPEians,

Please find below the summary and action items from the RF3 meeting on October 27, 2022. If you have any questions about the meeting or action items please reach out to Dr. Melanie Ott and Dr. Priti Kumar. 

Updates:

Priti Kumar shared that her lab has started some spironolactone experiments with mice, in collaboration with the Valente lab about using pellets. In a couple of months, they will look at rebound for the silencing experiments. She has some CURE infection data with ART (only looking at nanoluciferase). 

Micahel Corely discussed that they are running new single-cell studies. They are flash-freezing tissues from mice using liquid nitrogen. He shared that the brain is the hardest to get sequencing from. Nuclei from the tissues can be isolated and it looks like it is working. 

Ulrike Lange started to test the nanobody CD4 and found transfection is successful in T cells.  Martin started experimenting with transfecting cell lines compared to primary cells and found less transfection in the primary cell system. They have been producing new batches of the nanobodies and the first batch of purified CD4-Nanobody AB6 is to be made and sent to Priti.  Priti mentioned that promoter selection will be key to determining efficacy in Rhesus macaques versus mice.  Ulrike’s group is preparing a summary of cell line models of HIV latency to test Brec1 suppression of HIV.  Ulrike will initiate MTA with Lish’s group for material transfer.  Michael proposed an experiment where GFP+/- JLAT cells were sorted prior to BREC1 transfection to test the recombinase efficacy in active and “latent” cells.  Ulrike’s group is preparing a summary of cell line models of HIV latency to test Brec1 suppression of HIV.  Ulrike will initiate MTA with Lish’s group for material transfer.  Michael proposed an experiment where GFP+/- JLAT cells were sorted prior to BREC1 transfection to test the recombinase efficacy in active and “latent” cells.

Niren Murthy updated on his LNP work.  He mentioned that negatively charged lipids can increase spleen localization.  Although this is known in non-humanized mice this effect is unknown in humanized mice.  In collaboration with the Wang lab at UC-Davis, they put IV injected neg-LNP/CRE mRNA into adult Ai9 mice (n=3).  30 minutes post-injection, spleen cells were analyzed by flow cytometry. Transfection in mice with negatively charged lipids indicates CD3+ T cells are still transfected (7.9% & 8.3% of CD3+ T cells received and expressed the transgene). Lish asked about myeloid cell transfection.  CD11C+ cells did not receive/express the transgene.  Transfection of primary human T cells with aptamer targeting LNPs is being tested for gene delivery to primary human T cells.  Preliminary data show little toxicity. Although the two formulations only showed ~1% transfection) increased doses can be tried.

Melanie Ott and Francisco Belinchon shared information about his transfection experiment and there was not a high enough efficiency with Jurkat cells. Francisco completed Brec1 (BrecOff from Ulrike) transfection in JLATs using nucleofection.  ~0.2% transfection efficiency but no effect on HIV suppression was seen.  Repeats are being done with lentiviral vectors.  Niren mentioned Brec1 is finicky. Permanent expression may be necessary and higher levels of Brec1 are required for efficacy.

-Zoom call between Ulrike and Francisco is needed next week.  

-Western blots to verify Brec1 protein expression are needed.  

-Sorting cells may be one way to delineate the effect of BrecOff in HIV suppression in JLATs.

-Francisco is still optimizing/testing the Piggyback vector and CRISPRoff system, taken over from Martin.

 -Doxycycline inducible vectors to be sent to Francisco from Ulrike’s group (maps and construct).

Actions:

• Michael – Share mouse tissue flash freezing protocols with Priti (test materials sent)

• Niren – Send LNPs to Priti for the Kumar lab to test – Transfection of primary human t cells with aptamer target LNPs with heart tissues

• Ulrike meeting with Francisco to look at data from a previous student for Brec off suppression

  • Ulrike-Docs inducible vectors, prepping DNA, send maps and constructs to Francisco

  • Send new AV to Priti

• Lish – work on MTA for this (Brec, latency models), Ulrike to send MTA contact officer to Lish

Announcements:

• If you are attending the Miami Meeting and would like to join the HOPE dinner on 12/12 Please let Robert and me know. 

• The HOPE Collaboratory-wide meeting is on November 21st.

• The next RF3 Meeting is December 5th. Please come prepared with some updates. 

• Please Acknowledge HOPE & the NIH in any publications or presentations and notify the Program Manager. Here are some examples of what you can include. 

“Research reported in this publication was supported by the NIAID of the National Institutes of Health under award number UM1AI164559, with co-funding support from NIDA, NIMH, NHLBI, the NIDDK, and the NINDS. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.”

“This research was supported by NIAID award number UM1AI164559, co-funded by NHLBI, NIDA, NIMH, NINDS, and NIDDK.”

Hello HOPEians,

During the RF1 meeting on October 17th, 2022 we discussed the new RF/Collaboratory wide format and who would submit abstracts for the HIV Persistence meeting in Miami. Attached are the Management & Operations Updates.

HIV Persistence – International Workshop Miami

Abstracts are due TODAY (by 3pm PT/ 6pm ET)

Please send me your abstracts, so we can decide who will present during the flask talks. Here is a list of individuals that are submitting abstracts (please contact me if I missed someone):

• Julie Frouard, Zichong Li, Ulrike Lange, Ursula Schulze-Gahmen, Daniela Boehm, Luisa Mori, Sonia Mediouni Jablonski, Michael Corley (2x), Alina Pang, Chuan Li, Priti Kumar

A reminder that the abstracts that you submit will likely get published if chosen. You can have max 10 authors. CROI has some rules about presenting abstracts that have already been presented:

“If study data are accepted for publication or presentation after the abstract submission to CROI, and that publication or presentation is expected to take place before CROI, the presenting author must contact the conference manager at CROIabstracts@iasusa.org to provide details as soon as the presentation is accepted for review, presentation, or publication in another venue or outlet. Please be aware that, while publication in these circumstances will not necessarily prevent presentation of the research at CROI, our strong preference is that any additional publication or presentation happens either simultaneous to or following presentation at CROI. Requests to publish or present research accepted for presentation at CROI prior to its presentation at CROI will be evaluated on a case-by-case basis. Failure to notify the conference promptly regarding plans to publish or present a CROI-accepted abstract prior to CROI may result in the removal of the study from the conference program.”

Here is some language you can use to Acknowledge HOPE & the NIH:

“Research reported in this publication was supported by the NIAID of the National Institutes of Health under award number UM1AI164559, with co-funding support from NIDA, NIMH, NHLBI, the NIDDK, and the NINDS. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.”

“This research was supported by NIAID award number UM1AI164559, co-funded by NHLBI, NIDA, NIMH, NINDS, and NIDDK.”

New RF Structure

The RF meeting structure will change beginning in November.  For the rest of October, each RF will hold its monthly meeting.  In November, a larger group meeting will take place in lieu of the individual RF meetings.  These will alternate every month.

Hello HOPEians,

Please find below the action items from the RF2 meeting on August 29, 2022. If you have any questions about the meeting or action items please reach out to Dr. Nadia Roan and Dr. Lish Ndhlovu.

Updates:

• Lisa Henderson showed evidence that HIV transcription and translation can be observed in CSF samples from PLWH who are well-controlled on cART. The presence of these viral products is associated with neurocognitive impairment as measured by neuropsychiatric (NP) testing. They observed in vitro that targeting tat mRNA for degradation using ASOs is effective in HIV -relevant cell types (PBMC) and that knocking down tat can reduce HIV transcriptional activity and delay viral rebound; suggesting it is a viable candidate to induce ‘deep’ latency. Their next steps are to start ex-vivo experiments using patient-derived PBMCs to evaluate the effect on primary isolates. Also to start in vivo experiments in a humanized mouse model of HIV infection to evaluate the effect on tissue reservoirs and viral rebound (NIAID RAPIDD Program).

• Susana Valente explained that they solved the issue with Tat antibodies and shared dCA and SP combination treatment in J-Lat 10.6 cells using the western blot. For the cell lines, you need to block them with milk and BSA. There is difficulty detecting Tat in PBMCs and tissue samples. Currently, they are using humanized mice and monkey tissue samples. The Brazil team might be able to study this using human sample studies, and the Roan lab can test with tonsil samples.

Actions:

• Sara Gianella – Share ALS patient data and tissue availability with Avi Nath

  • Start a new MTA

  • Meeting with Sara Lemere, Avi Nath, Sara Weibel, and Lisa Henderson to discuss ALS & Endogenous Retrovirus reactivation in these HIV-positive participants.

• Sara Gianella – Collaborate with Avi Nath to mine data for defective proviral sequences

• Sara Gianella – Share morphine/opioid information on participant cohort with Lish Ndhlovu

Announcements:

• DUE TOMORROW: Call for nominations for the HOPE 2022 Awards (Instructions attached)

• Please Acknowledge HOPE & the NIH in any publications or presentations and notify the Program Manager. Here are some examples of what you can include.

“Research reported in this publication was supported by the NIAID of the National Institutes of Health under award number UM1AI164559, with co-funding support from NIDA, NIMH, NHLBI, the NIDDK, and the NINDS. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.”

“This research was supported by NIAID award number UM1AI164559, co-funded by NHLBI, NIDA, NIMH, NINDS, and NIDDK.”

Hello HOPEians,

Here is a quick update on the RF1 meeting from July 18th, 2022.

During the RF1 meeting, Cedric Feschotte (Cornell University) shared updates that Sabrina Leddy has started working in his lab on HERV expression in HIV elite controllers and Julius Judd who was previously working on zinc fingers is leaving his lab soon. Susana Valente (University of Florida) explained her research with DUX4, FUBP3, dCA-resistant viruses, and how p32 interacts with Tat’s basic domain.

Zichong Li (Greene Lab at Gladstone) discussed different epigenetic modifiers and the effects of new SPAs and the nucleosomal modification mechanisms for silencing HIV.  Martin Kinisu (Greene Lab) also shared his work on CRISPRoffV2 combining DNA and H3K9 methylation, the optimization of KRAB domain for HIV silencing, and exploring inducible expression systems using the piggyback technology. In his early findings, Martin found that ETRs transiently silence HIV reactivation in 30-70% of cells. Melanie Ott and Daniela Boehm (Gladstone) resubmitted their SYMD5 paper and performed the SMYD5 rescue experiment in 5A8 J-Lat cells. Their findings show that SYMD5 is highly expressed in CD4+ T cells and is a positive regulator of HIV transcription.  SYMD5 also methylates histones and Tat in vitro.

Actions for next meeting:

• Susana Valente – Schedule a meeting with Cedric to go over DUX4 data

• Chuan Li – Give MNase-Seq protocol to Zichong Li

• Roan Lab – Look for DUX4 expression in single-cell RNAseq tonsil datasets (+/- HIV)

• (In progress) Luisa Mori, Ana Leda, and Susana Valente considering writing a review/summary of all the HIV latency models

Announcements:

• August 15th HOPE – PAVE Guest Speaker: Dr. Persaud

• The next standing meeting is on Sept 19 – Bharath Sreekumar will give a refresher on their data on transducing T cells.  Cedric Feschotte and Sabrina Leddy will give updates on HERV expression in HIV-1 elite controllers.

• Call for nominations for the HOPE 2022 Awards (Instructions attached)

• Please Acknowledge HOPE & the NIH in any publications or presentations and notify the Program Manager. Here are some examples of what you can include.

“Research reported in this publication was supported by the NIAID of the National Institutes of Health under award number UM1AI164559, with co-funding support from NIDA, NIMH, NHLBI, the NIDDK, and the NINDS. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.”

“This research was supported by NIAID award number UM1AI164559, co-funded by NHLBI, NIDA, NIMH, NINDS, and NIDDK.”

Hello HOPEians,

Please find below the action items from the RF2 meeting on July 6, 2022. If you have any questions about the meeting or action items please reach out to Dr. Nadia Roan and Dr. Lish Ndhlovu.

Updates:

• HIV Natural History Study collaboration / Trevor Crowell – New data from the exposure and timepoint definitions suggests that there are more participants than anticipated (~10) taking spironolactone and some with five or more timepoints. These longitudinal samples may include pre- and post-spironolactone administration.  Samples include PBMC but not tissues.  They are currently identifying control participants and will finalize the participant selection soon.  The MTA agreements for both Susana Valente (University of Florida) and Lish Ndhlovu (Weill Cornell) are undergoing final review and will be sent for signatures soon.

• Sara Gianella Weibel with the Last Gift Cohort gave updates – The Last Gift (NIH P01) was recently renewed until 2027. She also shared modified autopsy procedures (Rapid autopsy → collection biobanking → processing → assays). Their new assays focus on single cell resolution (VENI and VIDI project assays).  Shared lessons learned. They found heterogeneous localization of HIV DNA within the CNS and also in the periphery.  This variability was participant-specific; however, the higher levels of HIV DNA in the spinal cord stood out and will be investigated further.  HIV RNA localization studies are ongoing and so far do not correlate with HIV DNA localization.

• Esper Kallas & Lish Ndhlovu collaboration on tissue studies – Discussed studies for people taking spironolactone, goal is to use paired gut biopsy and peripheral blood analysis of HIV reservoir pre- and post- spironolactone treatment. First, they will look at spironolactone effects on general DNA transcription in HIV negative participants before they complete similar studies in HIV positive participants.  The IRB protocol has been submitted at USP and should have a response in approximately 30 days.  Initiation of the study can begin soon after IRB approval.

• Susana Valente shared data from Tat degraders that were synthesized (SR-4481 & SR-44863).  High concentrations of SR-4483 was used in a preliminary murine study, but titrations will need to be done due to the high toxicity.  MTAs are ongoing with Lish Ndhlovu at Weill Cornell and Melanie Ott at UCSF.  Susana’s group has tried to validate several additional ERCC3 (XPB) antibodies, since the previously used antibody is longer available. These new antibodies do not work for WB or IP.  The Valente lab is also continuing to study the development of viruses resistant to SP. She explained the analysis of virus potentially resistant to SP used alone from the single RhM study from Maurucio Martins.  New dCA production is still 2-3 months away.  Some of this dCA is earmarked for larger NHP studies.

• Lish shared links to:

  • Esper Kallas Oral Presentation for AIDS 2022, August 2, from 10:30-11:30 ET: Shake and bake: Promising strategies for HIV cure

  • Viral Infections & Inflammation Workshop 2022 Sep 8- Sep 9 2022

Actions:

• Sara Gianella – Share ALS patient data and tissue availability with Avi Nath

  • Start a new MTA

  • Meeting with Sara Lemere, Avi Nath, Sara Weibel, and Lisa Henderson to discuss ALS & Endogenous Retrovirus reactivation in these HIV positive participants.

• Sara Gianella – Collaborate with Avi Nath to mine data for defective proviral sequences

• Sara Gianella – Look for participants who are on spironolactone – inform Susana Valente and Nadia Roan

• Sara Gianella – Share morphine/opioid information on participant cohort with Lish Ndhlovu

Announcements:

• July 13 Guest Speaker: Dr. Mathias Lichterfeld

• The next meeting is on August 3, 2022 – Lisa Henderson will present on assays for measuring defective provirsus and HIV targeting ASOs

• If you are attending the AIDS 2022 in Montreal (July 29-August 2) please let Lish Ndhlovu and Sydney Norman know.

• Register for the HOPE Annual Meeting

• Call for nominations for the HOPE 2022 Awards (Instructions attached)

• Please Acknowledge HOPE & the NIH in any publications or presentations and notify the Program Manager. Here are some examples of what you can include.

“Research reported in this publication was supported by the NIAID of the National Institutes of Health under award number UM1AI164559, with co-funding support from NIDA, NIMH, NHLBI, the NIDDK, and the NINDS. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.”

“This research was supported by NIAID award number UM1AI164559, co-funded by NHLBI, NIDA, NIMH, NINDS, and NIDDK.”

Hello HOPEians,

In Monday’s meeting on June 6th, we heard data updates from the Chemnitz and Lange Labs. They explained the construction and functional characterization of an enzymatically dead Brec1 and the generation of different expression plasmids for brec1 and dBrec1. They also went over CD4-specific AAV2 vectors using nanobody capsid engineering. Priti gave a quick update on using CRISPR-BE (BE4) targeted to TAR to inhibit HIV; initial LTR reporter assays show reduced luciferase expression, next they will test using Ya-chi Ho’s clonal cell lines. Priti shared the paper “Efficient viral delivery of Cas9 into human safe harbor” and Nadia shared “Why drug delivery is the key to new medicines”.

There is a working group meeting on CRISPRoff efforts and relevant data on June 13th at 10am PT. Joining information is on the HOPE calendar.

Here are the updated actions from our last meeting. If you have any questions about the meeting or action items please reach out to Melanie and Priti. 

Actions:

• (In progress) Priti have a meeting with Ulrike 
  • Discuss CD7 nanobodies and adding RSV molecules to AAV for targeted delivery
• Jan – Generate a Brec1 construct using an SFFV promoter for Priti

Long-term Actions:

• (In progress) Priti – Provide mouse tissues to Lish for scRNA-seq
  • Lish to set up a pipeline for sequencing and analysis
  • Potentially going to Yale to do initial steps on site
• Priti- Continue to work on Brec1 delivery using a lenti or VLP based methods

Announcements:

• The July 4th meeting is canceled for the holiday
• The next meeting is scheduled for Aug 1st and we will host a HOPE – CRISPR for Cure Joint Meeting with Guest Speakers – Dr. Kamel Khalili and Dr. Tricia Burdo

Hello HOPEians,

Please find below the action items from the RF2 meeting on June 1st, 20222. If you have any questions about the meeting or action items please reach out to Dr. Nadia Roan and Dr. Lish Ndhlovu. 

Updates:

• Dr. Lish Ndhlovu gave updates that they are continuing to try to make dCA with chemists at Weill Cornell.
• Dr. Brian Agan from IDCRP is joining HOPE. Several protocols have been approved to receive samples from the NHS and additional samples have been identified in RV537 of interest to HOPE.
• Dr. Ulrike Lange has sent plasmids of different Brec1 constructs to Dr. Priti Kumar and Dr. Melanie Ott. Dr. Ulrike Lange updated about the dBrec ChIP; there was a lot of plasmid background so it was difficult to call peaks, however, it did look like there were several potential off-target sites in the genome, they will look to see if these sites contain the Brec1 recombination motif. 
• Dr. Ana Carolina Soares gave an update that they finished writing their pilot project for spironolactone and aim to submit for IRB approval by the end of this week. They bought antibodies for western blotting for XBP so they can perform once they have samples. 

Actions:

• Dr. Ulrike Lange – Send cloned CRISPR off construct to Dr. Melanie Ott
• Dr. Ana Carolina Soares to send the reference for antibody for XBP to Dr. Susana Valente and Dr. Luisa Mori
• Dr. Denise Hsu to set up a subgroup meeting to discuss a potential PK study on spironolactone in NHPs. (Valente lab, Dr. Denise Hsu and team, Dr. Mauricio Martins, and Dr. Mike Cameron)
• Dr. Susana Valente to invite Dr. Mario Stevenson to give a talk about new myeloid cell paper
• (In progress) Approving and obtaining MTAs from Valente Lab (Dr. Ana Leda – MTA coordinator) 
  • Adding dCA to the MTA and getting Dr. Nadia Roan an MTA
• Dr. Susana Valente to coordinate sending DNA from dCA treated primary cells to Dr. Michael Corley to look at DNA methylation

Announcements: 

• Dr. Melanie Ott and Dr. Ulrike Lange will be hosting a CRISPR-off Efforts and Relevant Data subgroup meeting on Monday, June 13th, 10-11am PT. If you are interested and would like to join, the information is on the HOPE Calendar.
• Dr. Lish Ndhlovu shared the Department of Health and Human Services NIH Grant RFA
• The next meeting is on July 6th. 

All – If you have not already, please complete the HOPE Member Introduction form

Hello HOPEians,

Here is a quick update on the RF1 meeting from 5/16.  

During the RF1 meeting, Sonia Jablonski, a Staff Scientist in the Valente lab discussed transcriptional crosstalk between DUX4-FL and HIV-1. Martin Kinisu from the Greene lab discussed recent work on vetting CRISPR-off (7kb) as a latency promoting factor and testing variable KRAB domains. Zichong Li from the Greene lab discussed three factors that can play a role in the efforts to block & lock HIV outside of the 30 synergies identified among 10 silencing factors. Bharath Sreekumar from the Ott Lab gave an update on transducing t-cells. Thomas Premeaux shared information from the Johnathan Karn paper on The Glucocorticoid Receptor Is a Critical Regulator of HIV Latency in Human Microglial Cells. Melanie led a discussion of new ETRs including BRECiB.

 Actions for next meeting:

• Cedric – Share data on ZNF579 with Susana
• Roan lab – look for DUX4 expression in single-cell RNAseq tonsil datasets (+/- HIV)

Long-Term Actions:

• Luisa Mori, Ana Leda, and Susana considering writing a review/summary of all the HIV latency models 
• (T-TRACE) Discuss whether it would make sense to apply T-TRACE when we test block-and-lock agents in humanized mice 
• (Systems to test spironolactone in, and which concentrations) Roan lab to collect samples to look at XPB and send to Valente Lab
• (Aim 2) Keep the discussion open on the use of PICh or other methods

At the next meeting on June 29th, we will have a guest speaker, Dr. Mathias Lichterfeld.

Hello HOPEians,

Please find below the action items from the RF2 meeting on May 4th, 2022. If you have any questions about the meeting or action items please reach out to Nadia and Lish. 

In today’s meeting we got updates from:

• Valente Lab– Susana presented updates on her new Tat inhibitors. They are working to identify analogs with improved properties as well as continuing to study their mode of action.
• Ndhlovu Lab – Micheal presented updates on their live-cell imaging-based reactivation system. They have added the HC69.5 microglia clone as a cell line they can test in. Additionally, presented on new enzymatic methylation sequencing with a proviral capture method which showed good coverage of the proviral sequence.

Actions:

• Approving and obtaining MTAs from Valente Lab (Ana Leda – MTA coordinator) 
  • Adding dCA to the MTA and adding Nadia to Gladstone MTA
• Susana to coordinate sending DNA from dCA treated primary cells to Michael Corley to look at DNA methylation

Announcements: 

• The next meeting is on June 1st. 
• The email hope-rf2-members@gladstone.ucsf.edu can be used to reach the whole RF2 group.
• All – If you have not already, please complete the HOPE Member Introduction form
• Share announcements and achievements for the HOPE Twitter or Website by filling out the Social Media Shoutout form. 
• All – Email your photo to sydney.norman@gladstone.ucsf.edu