CENTER OF EXCELLENCE IN IMMUNOLOGY

STEERING COMMITTEE MEETING

SUMMARY MINUTES

October 15, 2007

# CENTER OF EXCELLENCE IN IMMUNOLOGY

**STEERING COMMITTEE MEETING**

October 15, 2007

1:00--3:00 p.m.

**Bldg. 31, Room 3A11**

**Videoconference to Frederick, Bldg. 560, Room 11-61**

> **Members present:** Dr. Robert Wiltrout *(Chair)*
>
> Dr. Jay Berzofsky
>
> Dr. Jeffrey Lifson
>
> Dr. Diana Linnekin
>
> Dr. Joost Oppenheim
>
> Dr. Marjorie Robert-Guroff
>
> Dr. Steven Rosenberg
>
> Dr. Giorgio Trinchieri
>
> Dr. Howard Young
>
> **Members absent:** Dr. Jonathan Ashwell
>
> Dr. Stephen Creekmore *(Adjunct)*
>
> Dr. Allan Hildesheim
>
> Dr. Ronald Gress
>
> Dr. Douglas Lowy
>
> Dr. Crystal Mackall
>
> Ms. Karen Maurey *(Adjunct)*
>
> Dr. Ira Pastan
>
> Dr. Jeffrey Schlom
>
> Dr. Alfred Singer
>
> Dr. Dinah Singer
>
> Dr. Robert Yarchoan
>
> Dr. Thomas Waldmann
>
> **Guests:** Dr. Rao Bachoti
>
> Mr. Ray Butler
>
> Dr. Scott Durum
>
> Dr. Barbara Felber
>
> Dr. George Pavlakis

######### Handouts

1.  Agenda

**Overview**

The Center of Excellence in Immunology Steering Committee (CEI SC)
convened its monthly meeting on October 15, 2007, in Building 31, Room
3A11 with videoconference available in Bldg. 560, Room 11-61, Frederick.
Dr. Robert Wiltrout called the meeting to order at 1:05 p.m. Dr. Giorgio
Trinchieri provided an update on the CEI-sponsored conference *Cancer
and Inflammation.* The committee decided to organize a meeting on the
theme *Effective New Cancer Immunotherapies.* Dr. George Pavlakis
provided an update on his project of DNA-mediated delivery of the
complex IL-15/ IL‑15Rα and presented data supporting funding for its
clinical development.

**Cancer and Inflammation Meeting**

Dr. Giorgio Trinchieri provided an update on the CEI-sponsored
conference *Cancer and Inflammation* held in Masur Auditorium on October
9--10, 2007. The meeting was well-received and very successful. Several
guests had words of praise and appreciation for all the hard work that
went into conduct of the meeting. It was felt that the meeting was very
useful and the topic was timely. There were a lot of good questions and
excellent feedback. About half the attendees were intramural scientists
and the rest were extramural scientists. Concern was expressed that
approximately half of the attendees did not attend the meeting's second
day; measures to encourage attendance at future meetings were briefly
discussed (e.g., charging a registration fee, and overbooking after
blocking certain rows for invitees and attendees coming from distant
places). The committee noted that some attendees would be from as far as
Europe. It was also felt that several slides were unreadable as too much
information was included on each slide. It was suggested that invited
and intramural speakers should be briefed about putting minimal
information on each slide. Overall, the CEI SC expressed great
satisfaction about the *Cancer and Inflammation Meeting* and voiced
appreciation for Dr. Trinchieri's efforts.

**Translational Immunology Meeting**

Dr. Wiltrout asked if the committee is ready to conduct a Translational
Immunology Meeting in the fall, as it has been three years since a
meeting on this topic has been held. Dr. Steven Rosenberg suggested
*Effective New Cancer Immunotherapies* as the title and volunteered to
organize the meeting. Dr. Diana Linnekin will help in organizing the
event and Dr. Jay Berzofsky offered to be on the Program Committee.

**Update on the IL-15 Project -- Dr. George Pavlakis**

Dr. Pavlakis provided an update on his project of DNA-mediated delivery
of IL-15/ IL‑15Rα complex and submitted a proposal for its clinical
development. Dr. Pavlakis explained that natural expression of IL-15 is
inefficient and tightly controlled at various levels. Higher levels of
expression are needed for clinical applications of IL-15; one method of
achieving high expression is coexpression of IL-15 with its receptor,
IL‑15Rα. 

Dr. Pavlakis mentioned that when the DNA vector capable of expressing
full-length macaque IL-15 and IL‑15Rα was injected into macaques, IL-15
plasma concentrations of up to 150 pg/mL were achieved. The proteins
were not immunogenic and sensitive Western blots indicated that
antibodies to the proteins were not produced. Immunogenicity was not
expected, as the proteins produced were likely similar to native
proteins.

Previous experiments in mice suggested that much higher levels of the
complex could be achieved if soluble IL‑15Rα (sIL‑15Rα)---a form of
IL-15Rα that is devoid of the membrane anchoring portion---was expressed
with IL-15 instead of full-length IL-15Rα. sIL‑15Rα is naturally
produced in the body and thus is not expected to be immunogenic. Dr.
Pavlakis and colleagues injected five macaques with plasmids encoding
IL-15 and sIL‑15Rα and observed IL-15 expression an order of magnitude
higher than that achieved with the full-length receptor. The IL-15 and
the IL-15Rα form a 1:1 complex, and in mice IL-15 levels and biological
activity correlated very well. The complexed state of IL-15 does not
affect the assay, as the antibody used does not distinguish between free
IL-15 and the complex. Dr. Pavlakis mentioned that some mouse IL-15
measurements in the literature may be questionable because some
antibodies against mouse IL-15 cannot recognize IL-15 that is bound to
IL-15Rα. He also mentioned that circulating IL-15 in humans is likely in
the complex form and that he would do some experiments to establish the
complex nature. The half-life of IL-15 alone is 40--70 minutes while
that of the complex is an order of magnitude higher.

Dr. Pavlakis studied the expression of IL-15/ sIL‑15Rα in macaques upon
delivery of the plasmid DNA via intramuscular route by electroporation
and observed levels of expression as high as 500 pg/mL in some animals.
He drew attention to the fact that all the macaques had higher plasma
IL-15 levels than the 50‑60 pg/mL usually observed in lympho-ablated
humans. He showed data from mouse experiments that achieved high levels
of expression of IL-15 locally in the muscle and calculated that 0.2% of
the complex produced in the muscle would eventually translocate to the
plasma when full length receptor was expressed; plasma levels of the
complex increased to 2% of the complex produced in the muscle when
soluble receptor was expressed. Extrapolating those results to humans,
he predicts that local expression of IL-15 in the muscle could be as
high as 30,000 pg when the soluble receptor is used.

Dr. Pavlakis proposed a clinical trial in which the IL-15/sIL-15Rα 
plasmid would be administered intratumorally. IL-15 should be
overproduced locally as shown by data on intramuscular injection by
electroporation in macaques. In mice, hydrodynamic administration of
IL-15/sIL-15Rα  plasmid one day before injection of tumor cells caused a
50% reduction in size of B16 melanoma tumor. Direct DNA delivery by
electroporation is an emerging technology. Two clinical trials employing
intratumoral DNA delivery by electroporation in melanoma are in
progress, one for IL-12 at South Florida University (Genetronics) and
one for IL-2 at Vical, San Diego, CA; both studies showed that there
were no toxic effects when DNA was delivered locally by electroporation.
Dr. Pavlakis has a CRADA with Genetronics. He briefly described the
electroporation device used in those studies. The device has been used
in humans and does not cause any tissue damage or other adverse effects
except mild local inflammation, which eventually disappears. Dr.
Pavlakis recapitulated that plasmids expressing both full length and
soluble receptor were developed, sequenced, and tested in mice and
macaques. He presented a comparison of cost estimates for GMP production
of vectors provided by different vendors and for carrying out toxicology
and macaque DNA integration studies.

The CEI Steering Committee suggested that development of any toxic shock
syndrome and induction of other inflammatory cytokines should be
monitored. Dr. Pavlakis said that IL‑15 plasma levels of 1 µg/mL in mice
were not fatal and that monitoring levels of other cytokines is in
planning stages. The committee also felt that: 1) administering IL-15
prior to injection of tumor cells was not the ideal experimental design
and 2) intratumoral injection is not the optimal route of delivery
because there is no animal data on intratumoral injection of plasmid DNA
by electroporation. Overall, the committee expressed some enthusiasm for
the project and noted that the cost of conducting the clinical trial is
not prohibitive. However, the committee felt that the experimental
system should be more analogous to the clinical setting. The committee
strongly advised that mouse data on intratumoral injection by
electroporation be obtained before proceeding to clinical application.
The alternative approach suggested by the committee was to do
intramuscular injection by electroporation in humans. The committee
would be interested in receiving an update on data collected using
intratumoral injection in an animal model or a presentation of a
clinical approach based on the existing intramuscular injection data.

The meeting adjourned at 3:00 p.m.

**Next Meeting: November 19, 2007**

**Location: Building 31/Room 3A11**

**Videoconference to Frederick: Building 549 Conference Room B**