Context

Molecular biology and immunology fields are witnessing their golden age due to the unprecedented advancement in high dimensional techniques that led to an important breakthroughs and treatments, taking the rapid development of coronavirus vaccines as an example.
In Algeria, the gap between theory and practice has always been a challenge in science, and the reasons for this vary, the most common is the lack of lab facilities and equipment especially in the universities, which is now compounded by the COVID-19 pandemic that has affected many in-person laboratory courses and on international student mobility and exchange opportunities.
To reduce “theory-practice gap”, the need to give students a deeper understanding of a concept by creating an appropriate context is important. For that, the virtual lab workshop which is collaboration between immuNOfrontiers organization, Blida-1 University and Miriam Merad’s laboratory, at Icahn School of medicine at Mount Sinai, in New York City, allows to train young Algerian researchers to solve a scientific question following experimental approaches without stepping into a physical laboratory. In this virtual lab the experiments performed are provided as an audio and video streaming of an actual lab experiment, it is an introduction for Algerian students to scientific research aimed at teaching them how to raise and respond to a research question, which appropriate experimental approach to choose, and how to interpret the obtained results. As a perspective, techniques learnt by the young scientists could be translate in the future to develop new research projects.
This virtual lab will cover a main subject on tumor immunity “Determining the effect of IL-4 blockade in tumor immunity”, and an additional subject on molecular biology “Shedding the light on the principle, functioning and the relevance of recombinant-based strategies for designing potent targeted cancer immunotherapies”.

Learning objectives

• – Learn how to answer scientific questions following experimental approaches;
• – Learn common techniques used in cancer-immunology research;
• – Learn how interpret and analyze data.

Participants

• – Undergraduate
• – Graduate students
• – Lecturer
• – researchers

Targeted disciplines

• – Biology
• – Pharmacy
• – Medicine.

Speakers

The main speakers of the workshop come from the laboratory of Dr Miriam MERAD, Icahn School of Medicine at Mount Sinai, USA, and Blida-1 University.

About the workshop

• – Language: English
• – Workplace: The workshop will take place at Blida’s University, and online in live streaming.
• – Date: September 27, 28, 29, 2022.
• – Time (Algeria Timezone): 14h – 18h
• – Time (New York Timezone): 9 AM – 1 PM
• – Duration: 3 days

Honorary Chairmans

• – Dr. MERAD Miriam (Director of the Precision Immunology Institute and Director of the Mount Sinai Human Immune Monitoring Center, Icahn School of Medicine at Mount Sinai, USA)
• – Pr. BEZZINA Mohamed (Rector of Saad Dahlad Blida 1 University, Algeria)
• – Pr. MEGATLI Smail (Dean of the Faculty of Natural Sciences and life, Saad Dahlad Blida 1 University, Algeria)

Organizing committee

• – BELABED Meriem (Icahn School of Medicine at Mount Sinai, USA)
• – AISSANI Billel (Blida 1 University, Algeria)
• – CHAMI Selma (Blida 1 University, Algeria)
• – SAADI Leila (Blida 1 University, Algeria)
• – CHALAL Nour El Houda (Blida 1 University, Algeria)
• – ARKAM Fatima (Blida 1 University, Algeria)
• – GAJARDO-CARRASCO Tania (Imagine Institute, France)
• – PARK Matthew (Icahn School of Medicine at Mount Sinai, USA)
• – ASSAD Nima (Icahn School of Medicine at Mount Sinai, USA)

Scientific committee

• – BELABED Meriem (Icahn School of Medicine at Mount Sinai, USA)
• – AISSANI Billel (Blida 1 University, Algeria)
• – CHAMI Selma (Blida 1 University, Algeria)
• – SAADI Leila (Blida 1 University, Algeria)
• – CHALAL Nour El Houda (Blida 1 University, Algeria)
• – ARKAM Fatima (Blida 1 University, Algeria)
• – GAJARDO-CARRASCO Tania (Imagine Institute, France)
• – PARK Matthew (Icahn School of Medicine at Mount Sinai, USA)
• – ASSAD Nima (Icahn School of Medicine at Mount Sinai, USA)
• – MATEUS-TIQUE Jaime (Icahn School of Medicine at Mount Sinai, USA)
• – MATTIUZ Raphael (Icahn School of Medicine at Mount Sinai, USA)
• – LAMARCHE Nelson (Icahn School of Medicine at Mount Sinai, USA)
• – HEGDE Samarth (Icahn School of Medicine at Mount Sinai, USA)
• – TRONCOSO Leanna (Icahn School of Medicine at Mount Sinai, USA)
• – WILK Matthias (Icahn School of Medicine at Mount Sinai, USA)
• – MERAND Raphael (Icahn School of Medicine at Mount Sinai, USA)

Main project: Determining the effect of IL-4 blockade in tumor immunity

Context and objectives

Extraordinary advances have been made in the treatment of cancer by using immune checkpoint blockade, and several immune molecules are currently targeted. However, responses to this form of therapy are not universal, and insights are clearly needed to identify optimal biomarkers of response and to combat mechanisms of therapeutic resistance. Dendritic cells (DC), and DC1 in particular, play a fundamental role in limiting cancer growth. DC1 can take up exogenous antigen (Ag), including tumor Ag, and load it onto MHC-I and prime CD8+ T cells, a process called cross-presentation, which makes them necessary for sustaining a robust anti-tumor immune response. Cross-presentation of tumor Ag by DC1 is also critical for response to programmed cell death 1 (PD1) blockade. This is because exhausted T cells need to be re-activated by co- stimulatory molecules, along with tumor Ag, which are provided by cross-presenting DC. The importance of DC in tumor immunity has made them a major clinical target for cancer therapy. Miriam Merad’s team has been working for over a decade to develop strategies to harness DC. However, while there has been some clinical success in exploiting DC for cancer therapy, we still lack an effective clinical means to promote tumor-Ag charged DC to drive anti-tumor immunity. One reason for this, supported by a recent Merad lab’s study showing that tumor DC that take up Ag turn on an immune dampening program (mregDC) and subvert immunity and promote tolerance to tumor Ag. Indeed, this program includes the upregulation of maturation genes associated with T cell simulation (CD40, CD80, etc), but also genes associated with immunoregulation (PD-L1, IL4R). DC in this molecular state are named mreg DC. Strikingly, this module was strongly associated with Interleukine 4 (IL-4) inducing molecules such as IL-4 receptor subunit alpha (IL-4Ra). The aim of this workshop is to identify the effect of IL-4 blockade on enhancing DC immunogenicity and T cell effector response and whether that blocking could result in tumor growth reduction.

Additional project: Shedding the light on the principle, functioning and the relevance of recombinant-based strategies for designing potent targeted cancer immunotherapies.

Context and objectives

Recent breakthroughs in genetic engineering have led to the development of a promising approach to tackle cancer known as immunotherapy. This last relies on a wide range of therapeutic strategies based on recombinant products and/or engineered cells that play two main roles: the stimulation of the immune system in one hand and on the other hand, the reversal of the tolerance that is provoked by cancer cells.

1. 1. The talk of this session will mainly focus on the pros and cons of past and recently emerged assembly techniques and expression systems used for anti-cancer theurapeutics as well as on those currently under investigation for their clinical performance for the sake of improving, supplementing or replacing conventional anti-cancer methods. In addition to the aforementioned, future perspectives on this topic, will be discussed.
2. 2. The experimentation training of this session will explore how cloning can be used to study tumor-immune cells interactions. Indeed, during the last decade, there has been a renewed focus on how distinct mutations in the tumor drive differential remodeling of the tumor immune micro-environment. In order to dissect the effect of each mutation, one can introduce a mutation of interest (i.e. oncogene) into tumor cells using lentiviral vectors carrying the mutated copy of the gene. Transduced tumor cells with the lentiviral vectors can be selected for further propagation and eventually implantation into live mice—where we can begin to answer the exciting question of tumor-immune interactions.

Program

Day 1 (mregDC program identification) : (14h – 18h DZ) | (9 am – 1 pm NY)

Lectures

• – Lecture on innate immunity (tissue-specific).
• – Lecture on dendritic cells.

Hypothesis generation

Identifying additional molecular programs that may reduce dendritic cells functionality in vivo.

Skills training

Experimentation

1. 1. Be able to culture KP tumor cells
2. 2. KP Tumor cell injection in vivo

Analysis

1. 1. Sacrificing mice and harvesting tissues (focus on lungs and lymph nodes)
2. 2. Process of obtaining H&E images of tumors
3. 3. Tumor quantification (QuPath)
4. 4. Tumor associated DC sequencing.

Takeaways (conceptual)

• – The mregDC transcriptional program is induced upon capture of apoptotic tumor cells.

Day 2 (uncoupling the mregDC program by looking at targets) : (14h – 18h DZ) | (9 am – 1 pm NY)

Lecture

• – Lecture on type II immunity and IL-4 signaling.

Hypothesis generation

Blocking IL-4 in vivo to test our research question: Does interfering with a component of the mregDC program enhance anti-tumor immunity?

Skills training

Experimentation

1. 1. Tumor quantification upon IL-4 blockade
2. 2. Digesting and processing lung and tdLN tissues
3. 3. Staining cell suspensions for flow cytometric analysis

Analysis

1. 1. Acquiring recordings of cells based on flow cytometry
2. 2. Analysis of flow cytometric data using FlowJo (cell proportions and MFI)

Takeaways (conceptual)

• – IL-4 blockade modulates mregDC phenotype and has broader consequences for T cell immunity against lung tumors.

Discussion

What next? What you propose as the next line of work, based on the findings we’ve illustrated through the technical steps outlined above?

Day 3 (a specific technique to learn): (14h – 18h DZ) | (9 am – 1 pm NY)

Lecture

• – Shedding the light on the principle, functioning and the relevance of recombinant-based strategies for designing potent targeted cancer immunotherapies

Hypothesis generation

Cloning can be used to study tumor-immune cells interactions.

Skills training

Experimentation

1. 1. Obtaining the cDNA corresponding to the protein of interest
2. 2. Amplification of this DNA by PCR (primers selection)
3. 3. Cloning into an expression vector
4. 4. Transformation/transfection in a host cell

Analysis

1. 1. Selection of recombinant cells carrying the gene of interest

Takeaways (conceptual)

• – An early fundamental molecular biology technique such as molecular cloning is still relevant and widely used today to study tumor-immune cells interactions.

Closing statements and remarks from the organizers