Cell transfection apparatus and method

An apparatus and associated method provides for the application of a cell treatment agent, such as genetic material or drugs to be inserted within the cells of a patient in vivo. The apparatus may be a catheter arrangement with various embodiments for applying heat to a patient's cells in vivo in order to improve transfection efficiency or application efficiency. Laser beams may be applied directly to the cells. Alternately, the cells may be heated by electrical heating, chemical heating, radio frequency heating, microwave heating, infrared heating, ultrasound heating, or indirect laser heating. Further, the treatment agent may be heated prior to its application to the patient such that the treatment agent heats the cells of the patient.

BACKGROUND OF THE INVENTION 
This invention relates to a cell treatment apparatus and method. More 
specifically, this invention relates to a treatment apparatus and method 
for transfecting a patient's cells in vivo. 
Various techniques have been used, at least experimentally, for 
transfection of cells (i.e., insertion of new genetic material into the 
DNA structure of cells). 
One technique for transfection of cells has used laser poration. This 
approach has been performed in vitro using a laser beam to porate a single 
cell at a time under a specially adapted microscope. The microscope allows 
the direct puncture of the cell membrane in the presence of the gene. 
Specifically, an operator directs the laser beam towards an individual 
cell and the puncture of the cell membrane allows genetic material on the 
same side as the cell to enter into the cell. This approach is labor 
intensive and not practical to use in vivo. This laser poration technique 
is described in the article by Tao et al. entitled "Direct Gene Transfer 
Into Human Cultured Cells Facilitated By Laser Micropuncture of Cell 
Membrane" in the Proceedings of the National Academy of Science in 1987; 
84:4180-4184. 
Other approaches to transfection of cells have included chemical methods or 
electrical poration used in a cell culture, but such methods have not been 
readily applicable in vivo. In other words, such methods may allow 
treatment of cells which have been removed from the patient, but do not 
allow treatment of cells remaining with the patient (human or animal). 
The Nabel et al. article entitled "Site-Specific Gene Expression in Vivo by 
Direct Gene Transfer Into the Arterial Wall" in Science in 1990; 
249(4974):1285-1288 discloses a technique for transfecting genes in vivo 
which has been used in the arteries of pigs. This technique uses a 
catheter with a dual balloon system at the tip of the catheter. The two 
balloons are inflated to create a temporary chamber which allows the 
exposure of the arterial wall to a viral transporting agent in solution. 
This has been used successfully to transfect the arterial wall with a 
DNA-plasmid having a viral carrier. However, this double balloon method 
requires 30 minutes to bathe the arterial wall with the DNA-plasmid to be 
effective. This is not feasible in certain applications such as in the 
coronary circulation. Moreover, the time required for such a technique to 
work may pose severe problems even at other locations within the arteries 
of an animal or human. 
The Lim et al. article entitled "Direct In Vivo Gene Transfer Into the 
Coronary and Peripheral Vasculatures of the Intact Dog" appearing in 
Circulation, volume 83, no. 6, June 1991, pages 2007-2011, discloses a 
technique where endothelial cells are removed from the test animal and 
then transfected prior to reintroduction into the animal. In addition to 
that in vitro technique, the article describes in vivo transfection of 
arteries of dogs using catheters placed in peripheral vessels of the dogs. 
Proximal and distal lumens of the vessels were occluded with removable 
ligatures. In somewhat similar fashion to the dual balloon system, a 
temporary chamber is established and a transfection solution is supplied 
into that temporary chamber within the vessels of the animal. The article 
describes allowing the transfection solution to remain in the vessel for 
one hour. 
In the two above incorporated by reference applications, the present 
inventor has disclosed cell treatment (more specifically cell 
transfection) of a patient's cells in vivo by use of a laser catheter. No 
admission is made or intended that these prior applications of the present 
inventor are necessarily prior art to the present application. However, it 
is noted that the present inventor has discovered the use of various 
additional techniques for in vivo transfection of a patient's cells. As 
used herein, in vivo shall refer to treatment of a patient's cells without 
removing the cells from the patient. Thus, in vivo treatment involves 
treatment of cells within or on the patient. 
OBJECTS AND SUMMARY OF THE INVENTION 
Accordingly, it is a primary object of the present invention to provide a 
new and improved cell transfection apparatus and method. 
A more specific object of the present invention is to provide for highly 
efficient cell transfection in vivo. 
A further object of the present invention is to provide for transfection of 
cells in vivo relatively quickly (i.e., the cells in vivo need not be 
exposed for such long periods that lengthy disruptions, such as blocking 
of artery flow, are required). 
The above and other objects which will become more apparent as the 
description proceeds are realized by an apparatus for cell transfection 
including an instrument with a housing having a wall with at least one 
hole therein. A heater is operatively connected to the instrument so as to 
apply heat to cells of a patient in vivo. A source of treatment agent 
including a DNA plasmid is operatively connected to the instrument so as 
to apply treatment agent to the heated cells by way of a treatment channel 
inside the instrument and extending to the hole. The treatment agent is 
operable to transfect the heated cells. 
The heater, which may also be called a heating means, may be realized by 
various alternate constructions. In a first embodiment, the heater 
includes an optical fiber within the instrument for applying laser energy 
by way of the hole to heat the cells. In a second embodiment, the heater 
includes an optical fiber within the instrument for heating an opaque 
portion of the instrument such that the opaque portion in turn heats the 
cells. In a third embodiment, the heater is an RF source which heats the 
cells by application of radio frequency energy. In a fourth embodiment, 
the heater is a microwave source which heats the cells by application of 
microwave energy. In a fifth embodiment, the heater is an electrical 
heater mounted to the housing. In a sixth embodiment, the heater is 
chemical heating material adjacent the hole. In a seventh embodiment, the 
instrument has a distal portion adjacent the hole and a proximal portion 
remote from the hole and the heater is adjacent the proximal portion for 
heating the cells by heating treatment fluid remote from the distal 
portion. Other embodiments use infrared or ultrasound in order to heat 
patient cells. 
Regardless of the heater arrangement, the instrument is preferably a 
catheter for insertion in a patient. The catheter further includes a 
flushing solution channel terminating in a flushing solution exit for 
applying flushing solution to a treatment site in a patient. The catheter 
includes a balloon mounted thereon and a balloon channel connected to the 
balloon for inflating the balloon, the balloon serving to occlude a body 
passage when the catheter is used for cell transfection. 
The method of transfecting cells of a patient in vivo includes applying 
heat to the cells of the patient in vivo and providing a treatment agent 
including a DNA plasmid to the heated cells such that the heated cells are 
transfected. The application of heat is accomplished from one or more of 
the steps selected from the group including: application of laser energy 
to the cells, application of laser energy to heat an opaque portion of a 
part of an apparatus in thermal transfer position relative to the patient 
such that the apparatus in turn heats the cells, the application of radio 
frequency energy to the cells, application of microwave energy to the 
cells, electrically heating the cells, chemically heating the cells, 
heating the cells by infrared energy, heating the cells by ultrasound 
energy, and heating the treatment agent prior to its insertion in the 
patient such that the treatment agent heats the cells. 
Preferably, the method further includes inserting a catheter into the 
patient, the catheter having a treatment agent channel, and wherein the 
heat is applied by way of the catheter and the treatment agent is provided 
by way of the treatment agent channel. 
In one technique according to the present invention, heat is applied using 
application of laser energy exiting from the catheter in the form of a 
plurality of distinct beams including at least first and second beams 
which cause porations in the cells, the first beam applied to a first one 
of the cells when the second beam is applied to a second one of the cells. 
Heat may be applied by use of laser energy exiting from the catheter in the 
form of a beam leaving the catheter with a width of less than 100 microns. 
Heat may be applied by use of application of laser energy exiting from 
openings on a side of the catheter in the form of a plurality of beams, 
each leaving the catheter with a width of less than 100 microns, the 
plurality of beams causing porations in the cells, and the treatment agent 
is forced out the openings to follow multiple paths corresponding to the 
beams. 
Heat may be applied by application of laser energy to heat an opaque 
portion of a part of the catheter in thermal transfer position relative to 
the patient such that the catheter in turn heats the cells.

DETAILED DESCRIPTION 
Turning initially to FIG. 1, a brief overview of the preferred embodiment 
of the present invention will be given. A treatment catheter 10, which 
might also be called a catheter assembly, includes an insertion catheter 
12 and a laser catheter 14. The insertion catheter 12 has a balloon 16 
mounted thereon for blocking blood flow in an artery, while the laser 
catheter 14 is applying laser energy 18 to porate cells (not shown) at the 
same time as genetic code material (not separately shown) is proceeding 
along the same paths as the laser energy 18. A guide wire 20 is used to 
guide the catheters to their intended location such that the laser energy 
18 and associated genetic material may be applied to the proper site 
within the patient. 
A balloon syringe 22 controls the balloon 16 in known fashion by way of a Y 
connection 24, which may be constructed in known fashion. Saline or other 
flush may be provided to the insertion catheter 12 by way of Y connector 
26 having entry tube 28 for those purposes. The laser or inner catheter 14 
is visible as it extends out the back of connector 26 towards the Y 
connector 30 having tube 32 for entry of genetic material or a drug to be 
inserted in the patient. A pump (not shown) or other arrangement may be 
connected to tube 32 to supply genetic material or a drug under pressure 
to inner catheter 14. The Y connector 30 is constructed in known fashion 
to merge the guide wire 20 (which proceeds from the luer lock or 
hemostatic Y-connector 34 and tube 36), optical fiber 38 (operably 
connected to laser 40 and proceeding through fiber connector 42 and tube 
44), and the material inserted into the entry tube 32. Accordingly, the 
inner or laser catheter 14 proceeding out the left side of connector 30 
includes the guide wire 20, the optical fiber (fiber optic element) 38, 
and any material supplied to the tube 32. 
Turning now to FIG. 2, the details of the tip of the catheter arrangement 
of FIG. 1 will be explained. The insertion of flush catheter 12, which may 
be made of a common catheter material such as Teflon type material, is 
generally a hollow cylinder having a balloon control channel 46 separated 
from the main hollow part of the tubular catheter 12 and used to control 
the balloon 16 in known fashion by way of an exit 46E for the channel 46. 
A flushing solution channel 48, which is separate and distinct from the 
balloon channel 46, is disposed within the main hollow part of catheter 
12. More specifically, the channel 48 may extend circumferentially in a 
ring just inside the wall of catheter 12 and outside of the inner or laser 
catheter 14. The channel 48 proceeds to a circumferential flush exit 48E. 
The inner or laser catheter 14 has a generally cylindrical tube 50 of 
common flexible material used for catheters. Inside of the tube 50 is a 
guide wire tube 52 for slidably receiving the guide wire 20 therein. The 
tube 52, would be secured to one side of the tube 50 as best understood by 
momentarily turning to the end view of laser catheter 14 in FIG. 2A. 
Instead of having the tube 52 be distinct from tube 50, tube 52 might 
simply be a lengthwise extending pocket in tube 50 extruded at the same 
time as tube 50. 
Turning back to FIG. 2, the optical fiber 38 extends inside of tube 50 and 
outside of tube 52 and extends into a connecting tube 54, which is 
preferably about 4 millimeters long and made of metal. The guide wire tube 
52 and the connecting tube 54 would be glued, have barbs (not shown) to 
grip tube 50, or otherwise fixed in position (possibly simply by friction) 
within the tube 50. The connecting tube 54 secures a tip 56 to the tube 
50. Specifically, the connecting tube 54 may be glued, friction-fit, snap 
fit using a ledge ring (not shown), or otherwise fixed to a cylindrical 
bore 56B within the tip 56. A second bore or cylindrical hole 56R (turn 
momentarily to FIG. 3) extends lengthwise in the tip 56. The tip 56, which 
is preferably made of surgical steel and is 8.7 millimeters long in the 
preferred embodiment, has a tapered end 58 which may be made of Teflon or 
other surgical materials commonly used in catheters. The tapered portion 
58 would be hollow or otherwise allow for passage of guide wire 20 
therethrough. 
The optical fiber 38 proceeds through connecting tube 54 to a glass hood 
60. Since the optical fiber 38 has a smaller outside diameter than the 
inner diameter of the hollow cylindrical connecting tube 54, an annular 
passage 54P is provided therebetween to allow fluid flow of treatment 
agent from an annular channel 53 into bore 56B. The optical fiber 38 is 
secured to the glass hood 60 by way of epoxy 62 applied after heating and 
creating a vacuum within the hood 60. The hood 60 preferably has a square 
cross section to fit within a square cross section cavity 60C (also refer 
momentarily to FIG. 3) which extends out from the circular bore or cavity 
56B. Accordingly, the glass hood 60 may be adhered, friction-fit, or 
otherwise fixed within cavity 60C to prevent relative angular movement 
between the optical fiber 38 and the tip 56. The optical fiber 38 may have 
a portion of its cladding removed at its narrow portion 38N adjacent its 
end. The optical fiber 38 has a tip 38T to cause any laser beam to be 
directed sideways out a window portion 56W in the side of the wall of tip 
56. More details of the construction of tip 38T and glass hood 60 may be 
obtained from U.S. Pat. No. 5,061,265, invented by the present inventor 
together with Stephan E. Friedl, issued on Oct. 29, 1991, and hereby 
incorporated by reference. Generally, the tip 38T is made into a prism 
using techniques described in that prior patent so as to deflect all, or 
substantially all, of the laser energy out the window portion 56W. 
Continuing to view FIG. 2, but also referring to the enlarged view of the 
window portion 56W of tip 56 appearing in FIG. 4, the window portion 56W 
is essentially a laser screen having a plurality of very small holes 56H 
through which laser micro-beams 64 may pass. Thus, the window portion 56W 
may be considered as a grating portion having grating means, the grating 
means constituted by the holes or openings 56H and the material in between 
the holes 56H. The holes 56H would be distributed throughout an area of 
between one square millimeter and three square millimeters. Preferably, 
the holes are distributed evenly over a circular area of two square 
millimeters which would correspond to the width of the beam exiting from 
the tip 38T. Each of the holes 56H would be less than 200 microns. More 
specifically, the holes would be below 100 microns in diameter such that 
the micro-beams 64 would have a corresponding width as they leave the tip 
56 of catheter 14. Most specifically, the holes would be 50 to 100 microns 
in diameter to provide beams of the same size. The holes 56H may be made 
by using an excimer laser or electrodischarge machine. The microbeams 64 
would preferably have a diameter of 50 to 75 microns. As apparent from 
FIGS. 2 and 4, there are at least three beams which go in the same general 
direction. More specifically, at least 12 beams, corresponding to at least 
12 holes, proceed out one side of the tip 56. As will be appreciated, 
there is a one-to-one correspondence between the beams 64 and the holes 
56H. The beams 64 are distinct (i.e., meaning distinct where they leave 
the tip 56), but each of the beams 64 diverges somewhat because of the 
properties of the prism tip 38T of the optical fiber 38. 
The openings 56H also allow passage of treatment agent which is supplied 
via treatment agent channel 53 (FIG. 2) in between fiber 38 and tube 50, 
connecting tube 54 (i.e., between tube 54 and fiber 38) and bore 56B. 
Operation 
Having described the structural features of the present invention, the 
method according to the present invention will now be described. 
Although the present invention has applicability to providing treatment 
very efficiently on a cellular level at various sites on a patient's body 
or in a patient's body, the specifics of the structure which has been 
described is best suited for applying treatment to the walls of an artery 
and the explanation which follows will emphasize such an application of 
the invention. 
A patient having an artery, such as a coronary artery, with atherosclerotic 
plaque is appropriately sedated and placed upon a x-ray or fluoroscopic 
table. Various know steps could be used for locating the tip 56 of laser 
catheter 14 at the site for treatment of the patient's artery and only a 
basic discussion of the procedure for locating the tip 56 at the proper 
site will be presented herein. Initially, the guide wire 20 and a guide 
catheter of common design (not shown) would be inserted into the patient 
using an introducer sheath of common design (not shown). The guide 
catheter would extend to the mouth of the artery, whereas the guide wire 
20 would be manipulated to anchor its end just beyond the partial 
obstruction caused by the plaque. 
The treatment catheter 10 (refer to FIG. 1), which includes both the laser 
or inner catheter 14 and the insertion or flush catheter 12 would then be 
slid along the guide wire 20. The catheters 12 and 14 would move along the 
guide catheter (not shown) in known fashion until the balloon 16 of the 
insertion catheter 12 is outside of the guide catheter and until the 
window portion 56W is adjacent the portion of the artery for which 
treatment is intended. The laser catheter 14 would be rotated until its 
window portion 56W (refer to FIG. 2) faces the side of the artery wall 
which is to be treated. The balloon 16 is then inflated using the balloon 
syringe 22 such that the part of the artery downstream (it would 
correspond to the rightward direction in FIG. 1) of balloon 16 is blocked 
from receiving further blood. The balloon 16 may be used to block the 
blood flow for up to about 60 seconds. The blockage would normally not 
need to be maintained for 60 seconds, but the surgeon would be using his 
professional judgement as to how long the blockage might be tolerated for 
a particular individual. At any rate, the blockage would be less than two 
minutes at a time and is significantly less than the blockage times 
required for the double balloon prior art technique described in the 
background portion of this application. If advisable, the balloon could be 
deflated and re-inflated to provide repeated treatments without 
maintaining the blockage for longer than about 60 seconds each time. 
After the balloon 16 has been inflated and now considering FIGS. 1 and 2, a 
saline or other flushing solution is supplied to connector 26. The saline 
travels along the flushing solution channel 48 and exits from 48E (see 
especially FIG. 2) so as to clean out blood in the portion of the artery 
just downstream from the balloon 16. After this space has been flushed 
with saline, the saline flush is halted and the laser 40 is activated to 
generate the laser energy 18 (FIG. 1) in the form of the microbeams 64 
(FIG. 2). The laser 40 would preferably be a pulsed laser pulsed at one to 
five times a second, such as a 355 nanometer tripled YAG or a flash lamp 
excited dye laser at 504 nanometers. However, a continuous wave argon 
laser or other type of laser might be used. A treatment agent will be 
supplied to the cells which are porated by the laser microbeams. A 
treatment agent as used herein is a cell treatment agent, meaning that it 
has medicinal effect (might include killing the cell if that was medically 
helpful) or harmful effect (if desirable for testing purposes) or remedial 
effect when placed within a cell after passing through porations caused by 
the laser microbeams. The treatment agent, such as genetic material or a 
drug is supplied to the connector 30 (FIG. 1) and passes through the 
treatment agent channel 53 and through the space between optical fiber 38 
and connecting tube 54 into the hole or bore 56B for passage as a high 
pressure stream out of the window portion 56W. Referring now to FIG. 5, 
two cells 68 are shown having porations 68P therein as caused by the laser 
microbeams 64 having edges 64A shown in FIG. 5, the microbeams having 
passed out of holes 56H. Also passing out of the holes 56H is a high 
pressure solution containing the treatment agent 70 disposed therein and 
some of the treatment agent 70A has entered into the cells 68 by way of 
the porations 68P. If the agent 70 includes genetic materials, the 
solution may be culture material such as DMEM (Dulbecco's Modified Eagle's 
Medium) or normal saline. Quite importantly, the treatment agent 70 passes 
out of the same holes as the beams 64 such that the treatment agent passes 
along the same paths as the various microbeams 64. Accordingly, the 
genetic material should enter through the porations 68P in the cell walls 
of cells 68. In other words, the treatment agent is concentrated precisely 
where it is most likely to be effective. Significantly, the beam width is 
smaller than the size (i.e., longest dimension) of the cell and would also 
preferably be smaller than the normal dimension of the cell (i.e., the 
dimension of the cell extending perpendicular to the direction of the 
beam). 
It should be appreciated that some of the microbeams 64 may hit the nucleus 
of a cell and kill the cell. Others of the microbeams may hit the edge of 
a cell without providing a useful poration. However, using a large number 
of the microbeams should allow for treatment, on a cellular level, of a 
sufficient number of cells that benefits will be obtained. 
As an alternative to the simultaneous spraying of treatment agent 70 out of 
the holes 56H while the beams 64 are passing out of the holes, one might 
porate the cells 68 by application of the beams 64 and, immediately after 
turning off the beams 64, spray the treatment agent 70 out of the holes 
56H. Since the porations 68P will close relatively quickly, the treatment 
agent should be sprayed immediately after turn off of the beams. 
If one is simply treating a single part of the artery wall, one might line 
up the window portion 56W to face the proper direction by use of a marker 
(not shown) on part of the laser catheter 14. For example, a hole or 
pattern (not shown) might be placed on the side of steel tip 56 opposite 
to the window portion 56W. The surgeon would then observe the marker by 
use of the x-ray table and rotate the laser catheter 14 until the marker 
was opposite to the part of the artery wall which was to be treated. If 
desired, two markers of different configuration might be used to provide 
more information to the surgeon and to help better line up the window 
portion 56W such that it faces the part of the artery wall which is to be 
treated. Instead of placing the markers on the steel tip 56, the markers 
might alternately be placed upon the tapered portion 58 (which is made of 
plastic) and/or the outer surface of the laser catheter 14. If desired, 
one may treat the artery wall in a complete circumference. One may apply 
laser energy and treatment agent (either simultaneously or treatment agent 
immediately after laser as discussed above) with the laser catheter 14 
disposed in one angular position. The laser would then be turned off, the 
laser catheter 14 would be rotated to a different angular position and the 
laser and treatment agent application would be repeated. The laser would 
be turned off and the laser catheter 14 would be rotated to another 
angular position for treatment. This process of rotation treatment 
followed by further rotation and treatment may be performed around the 
complete circumference of a portion of the artery. Additionally, or 
alternately, one may move the laser catheter 14 along the guide wire 20 to 
a different place within the artery before applying further treatment. In 
other words, if the blockage or other problem in the artery extends 
significantly in a lengthwise direction, treatments may be applied at 
different places along the length of the blockage. 
Advantageously, a vacuum may be applied to tube 28 to remove genetic 
material or drug remaining free after laser operation and before deflating 
balloon 16. This reduces the amount of material going down stream. 
After application of the laser beams and treatment agent has been 
completed, the balloon 16 would be deflated so as to reopen the artery. 
The treatment catheter 10 composed of laser catheter 14 and insertion 
catheter 12, is then removed from the patient. The guide catheter and 
guide wire would be removed from the patient and normal post-operative 
procedures would be followed such as checking the patient to insure that 
no artery walls were punctured. 
Having shown how the present invention may be used to very efficiently 
provide treatment at a cellular level by injecting cell treatment agents 
70 directly into cells 68 as illustrated in FIG. 5, some specific examples 
of such treatments using laser energy will now be presented. Generally, 
any genetic code material, such as DNA plasmids, and any drug applicable 
for cellular treatment could be used. 
EXAMPLE 1 
A patient has a buildup of plaque on the walls of a coronary artery. The 
treatment catheter 10 would be inserted into the patient under the 
procedure explained above and the microbeams 64 are used to porate smooth 
muscle cells of the artery walls. The treatment agent would be plasmids of 
DNA which encode antisense gene. The gene may be under the control of 
mouse metallothionein promotor. A virus carrier would be used in known 
fashion to allow the desired genetic material to enter the nucleus and/or 
cytoplasm (most frequently the cytoplasm) of the cells into which the 
treatment agent is inserted. The antisense genetic code will fool the 
smooth muscle cells to inhibit growth patterns which cause the blockage of 
arteries. 
EXAMPLE 2 
A patient has a buildup of plaque on the coronary arteries. The treatment 
catheter 10 would be inserted into the patient under the procedure 
described above and the microbeams 64 are used to porate the endothelial 
cells of the artery walls. The treatment agent would be plasmids of DNA 
which encode a tissue plasminogen activator gene under the control of the 
mouse promotor as with example 1 and having a virus carrier. The human 
plasminogen activator would cause the production of enzymes which reduce 
formation of clots. 
EXAMPLE 3 
A patient has a malignant tumor in the colon. The patient would be sedated 
and a colonoscopy would be performed. Instead of using an x-ray or 
fluoroscopic table to locate the position of the treatment device placed 
within the patient, the treatment device (not shown) may include an 
optical fiber to allow the surgeon to see within the colon. The probe or 
medical device inserted into the patient would include a laser catheter 
similar to catheter 14 of FIG. 2. Upon the window portion (similar to 56W 
of FIG. 2) being lined up to face the tumor, laser energy is applied to 
provide microbeams which porate the cells of the tumor and a cancer agent, 
such as 5-fluorouracil or donarubicin, is injected out the same plurality 
of holes used for generating the microbeams. Saline or other solution may 
be used to carry the cancer agent drug to the cells. The cancer agent 
would enter into numerous of the cancer cells and kill them. 
Advantageously, the poration of the cells by the laser beam improves the 
efficiency of application of the cancer agent to the cells. A smaller 
portion of the cancer agent harms adjacent healthy cells than would be the 
case if one simply applied the cancer agent against cells which had not 
been porated. Some of the cancer cells may be killed simply by application 
of the laser beam, but the inclusion of the cancer agent helps to kill a 
greater portion of the cancer cells than would otherwise be the case. 
Additionally, if one simply relied upon the laser to kill cancer cells, 
one might have to use a higher laser power which in turn might damage 
healthy cells behind or adjacent to the cancer cells. 
EXAMPLE 4 
The patient would be the same as in example 2 and the same procedure would 
be followed except that the treatment agent is the drug heparin carried by 
saline or other solution. The drug prevents formation of clots. 
EXAMPLE 5 
An animal may be used to test various anti-plaque techniques by using the 
present invention to induce plaque in walls of arteries. The treatment 
catheter 10 is used to porate endothelial cells on artery walls of the 
animal (patient) for introducing a plasmid of DNA which encodes for human 
growth hormone gene under the control of a mouse metallothionein promoter. 
A viral carrier would be used in known fashion to allow the desired 
genetic material to enter the nucleus and/or cytoplasm of the cells into 
which the treatment agent is inserted. The expression of growth hormone in 
transfected cells will then result in the expression of various cellular 
proteins causing cell growth which could be responsible in part for the 
development of plaque in the arterial wall. This information could then be 
used to develop either drugs or other methods to inhibit gene expression 
in order to block this growth. 
Turning now to FIG. 6, an alternate embodiment constructed in somewhat 
similar fashion to the embodiment of FIG. 1 will be discussed. The 
components of the FIG. 6 embodiment have numbers in the 100 series with 
the same last two digits as the corresponding component, if any, of the 
FIG. 1 embodiment. Thus, the components 110 through 144 are identical in 
construction and operation as the corresponding components in FIG. 1 
except as discussed hereafter. 
As shown, a reservoir 172 of treatment agent with DNA plasmid is connected 
to pump 174. The pump 174 pumps the agent into entry tube 132 by way of 
tube 176. This operation as described so far is identical to the catheter 
10 of FIG. 1 where the reservoir and pump were simply not shown. What is 
different about the catheter system 110 is the use of an electrical 
heating coil 178 wrapped around the tube 176 in order to heat the agent 
prior to its insertion in the patient. By heating the agent sufficiently 
that the agent adjacent the patient's cells will be from 42 degrees to 45 
degrees, the transfection process is speeded up and will occur faster than 
would otherwise be the case. Indeed, the laser 140 and laser catheter 114 
could be left out of the apparatus 110 and the heat from electrical heater 
178 could be used without laser energy in order to expedite the 
transfection process. In that case, a catheter 114 could be constructed 
like catheter 14 of FIG. 2 except that there would be no optical fiber 
such as 38 of FIG. 2. Also, such an arrangement could use one of more 
large exit holes instead of the array of 50 to 100 micron diameter holes 
56H of FIG. 4. 
Although the heater 178 is shown as a coil heater, other electrical or 
non-electric heating techniques could be used to heat the agent either 
prior to its insertion in the catheter 110 or while it is traveling 
therein. 
FIG. 7 shows an embodiment using application of laser energy to heat the 
tip of catheter 210 and thereby indirectly heat the cells. The components 
of the FIG. 7 embodiment have numbers in the 200 series with the same last 
two digits as the corresponding component, if any, of the FIG. 1 
embodiment. Insertion catheter 212, laser catheter 214, optical fiber 238, 
flushing solution channel 248, flushing solution exit 248E, and treatment 
agent channel 253 function as the corresponding components in the previous 
embodiments except as discussed below. (The unseen parts of catheter 210 
would be connected to a laser and a source of treatment agent with DNA 
plasmid as with previous embodiments.) 
The FIG. 7 embodiment is different from the others in that it has an opaque 
metal cap 280 with neck portion 280N crimped or otherwise fixed to 
catheter 214. The metal may be similar to metal parts 50 in FIG. 4 and 70 
in FIG. 5 of the present inventor's U.S. Pat. No. 5,041,109, patent, 
issued Aug. 20, 1991, and hereby incorporated by reference in its 
entirety. Cap 280 has holes 280H for treatment agent to exit from, whereas 
optic fiber 238 terminates in a spherical lens 238S which distributes 
laser energy, not shown, over a wide area of cap 280. Arms 238A may be 
circumferentially arranged to support fiber 238. The laser energy heats 
the cap 280, which in turn would heat the patient's cells (not shown) and 
make them more receptive to transfection from the treatment agent. 
Although not shown in the FIG. 7 embodiment, one or more occluder balloons 
such as 16 of FIG. 1 could be used in the FIG. 7 embodiment and the 
embodiments hereafter discussed. For ease of illustration, the insertion 
catheter, such as 212 of FIG. 7, and flushing solution channel will not be 
shown or discussed for the embodiments discussed below, even though those 
embodiments would have such features. 
Catheter 310 of FIG. 8 has numbers in the 300 series with the same last two 
digits as the corresponding component, if any, of the earlier embodiments. 
This embodiment has a cap 380 with hole 380H for dispersing treatment 
agent from channel 353. A thermocouple or other temperature sensing device 
381 has wires 381W (only one shown for ease of illustration) extending out 
of catheter 310; The cap 380 has at least one heating resistor 382 
embedded therein or otherwise in thermal contact therewith. The resistor 
382 serves as a heat source. Wires 382W (only one shown for ease of 
illustration) carry electrical current from a power supply 382P external 
outside of the patient to the heater resistor 382. The power supply 382P 
is part of a feedback control closed loop which uses negative feedback by 
way of temperature adjustor resistor 381R and comparator 381C. As well 
known such a control loop could alternately use an amplifier, subtractor, 
or adder (instead of comparator 381C) to generate a difference signal at 
the output thereof to cause the heater resistor 382 to heat more if the 
temperature is low and to heat less if the temperature is too high. Since 
thermocouple or other sensor 381 is located near the hole 380, it will 
very accurately track the temperature of treatment agent fluid passing out 
the hole 380H. Preferably, the sensor 381 is within at least 5 inches of 
hole 380H, and more preferably and advantageously within 3 inches thereof. 
Most preferably, it is at the edge of hole 380H or within 1 inch of the 
edge of the hole. 
The feedback circuit may be configured differently from FIG. 8 as many 
types of thermostatic or other feedback techniques are known for 
stabilizing a sensed condition such as temperature. It should be 
emphasized that the feedback control to maintain the fluid temperature 
shown for FIG. 8 would preferably be used for the various other 
embodiments discussed above and below. Such feedback control loops and 
components are not shown for the other embodiments for ease of 
illustration. However, all embodiments may use a temperature dependent 
sensor on the catheter as discussed to control the output of the heating 
device or devices so as to maintain the temperature at a desired value or 
within a desired range, which is preferably 42 to 45 degrees centigrade. 
The heater resistor 382 may be of the CAL-ROD type or any other device 
generating heat from electricity. The cap 380 may be metal like cap 280 or 
could be other material suitable to disperse heat to patient's cells 
adjacent to the window 380H. As with the other embodiments not based on 
direct application of laser energy to the patient's cells, the heat could 
be used to heat the patient's cells to from 42 to 45 degrees centigrade 
and enhance their receptivity to transfection. 
Catheter 410 of FIG. 9 has numbers in the 400 series with the same last two 
digits as the corresponding component, if any, of the earlier embodiments. 
Inner catheter 414 has holes 480H to allow exit of treatment agent from 
treatment agent channel 353. Radio frequency (RF) waves are emitted from 
antenna 484 or other known RF source. Wires 484W are connected to supply 
power as appropriate to antenna 484, which is embedded in part 414E. Part 
414E may simply be a generally cylindrical rounded tip of the material 
making up the side of catheter 414 (and sides of the various other 
catheter embodiments), which material may be PTFE (such as Teflon) or 
other materials commonly used for catheters. The RF energy is used in this 
embodiment to heat the patient's cells, preferably to 42 to 45 degrees 
centigrade, in order to make them more receptive to transfection. 
The FIG. 10 embodiment of catheter 510 uses microwave energy from source 
586 outside the patient to heat cells for improving receptivity to 
transfection. Microwave source 586 supplies microwave energy to inner 
catheter 514 by way of waveguide 586G for exiting at a microwave output 
tip 586T. The microwave energy is applied to patient's cells (not shown) 
adjacent hole 580H from which treatment agent from channel 553 may exit. 
FIG. 11 shows catheter 610 with inner catheter 614 having an infrared 
source 688 connected by wires 688W to a power source (not shown) outside 
the patient. Infrared energy from source 688 heats treatment fluid and/or 
the patient's cells (not shown) as treatment agent from channel 653 exits 
hole 680H. 
The catheter 710 of FIG. 12 has an inner catheter 714 with a treatment 
agent channel 753 supplying treatment agent out hole 780H. In this 
embodiment a chemical compartment behind (right in FIG. 12) wall 790 is 
divided into top and bottom parts 790T and 790B by separation wall 790S. 
Parts 790T and 790B hold different chemicals which react to release heat 
when they mix upon control line 790C being used to slide separation wall 
790S leftwardly in FIG. 12. The heat heats patient cells to improve 
receptivity to transfection. 
FIG. 13 shows a catheter 810 with inner catheter 814 having hole 880H for 
releasing treatment agent to a site 792S within a patient's artery (or 
other body passage) 792A. In this arrangement the cells at site 792S are 
heated by application of ultrasound energy from transducers 794 on the 
skin 792K of the patient. 
Turning now to FIGS. 14-17, a further embodiment catheter 910 has a tip 956 
with tapered portion 958. A guide wire 920 is used and a glass hood 960 is 
used. The components of catheter 910 and other unshown components may be 
constructed and may operate as the corresponding component (same last two 
digits) from FIGS. 1-6. This discussion will proceed with differences 
between catheter 910 and catheter 10. 
Catheter 910 has a window 956W which is simply one large opening. Instead 
of relying on holes to produce microbeams of laser energy for cell 
poration as done in the embodiment of FIGS. 1-6, catheter 910 uses a 
bundle 995 having a sloped tip 995S and a plurality of optical fibers 938 
(only some numbered for use of illustration) with filler material 995C. At 
least the portions adjacent to slope 995S of the filler material should be 
clear such that beams 964 may emerge from the tips 938T (shown 
schematically in FIG. 14 only). Each tip 938T may provide a single 
corresponding beam 964, which beam may have dimensions and operate as 
discussed for each of the beams 64 of FIG. 2. As illustrated schematically 
in the front view of FIG. 14, the tips 938T and fibers 938 are staggered 
such that light from one tip 938T will not hit another of the tips 938T or 
fibers 938 on its way to the single opening window 956W and the patient 
tissue outside it. 
The laser energy at the tips 938T is reflected out the window 956W by 
either of two arrangements. A first way is to simply have the optical 
fiber tips 938T cut at the same angle (preferably between 35 and 55 
degrees, most preferably an angle from 42 to 48 degrees, with 45 degrees 
being the most preferred value) as the slope 995S. (The angle being 
measured relative to a horizontal line, not shown, perpendicular to the 
axes of the parallel optical fibers 938.) This first way has tips which 
reflect light such that it goes in the direction from a low end of the 
sloped face towards the high end of the sloped fiber tip. A second way is 
to have a prism such as shown and explained for the tip of FIG. 7 of the 
incorporated by reference U.S. Pat. No. 5,061,265. 
FIG. 17 shows the external end 995E of bundle 995 receiving a laser beam 
940B from a laser 940. Since the optical fibers are relatively small, they 
will produce a plurality of beams having the width ranges as discussed 
with respect to the embodiment of FIGS. 1-6. The optical fibers themselves 
may have diameters equal to the various ranges discussed with respect to 
the beam widths given above dimensions with respect to the embodiment of 
FIGS. 1-6. The optical fibers may be 80 microns in diameter. As in the 
other drawings, only a relatively small number of fibers 938 are shown, 
but it will be understood that the number of fibers could be just a few or 
a relatively large number. 
A further alternate to the arrangement of FIGS. 14-17 may be discussed with 
reference to FIG. 16 since such a further embodiment might be constructed 
as with FIGS. 14-17 and have essentially the same top view. The beam 
pattern of FIG. 16 might be realized by plurality of fibers with prisms at 
their tips and with all the tips ending in the same plane perpendicular to 
the axes of the fibers and the catheter. Further, such prisms could 
alternately direct the beams radially outward from a 5 central axis of a 
bundle such as 995S. 
Turning now to FIGS. 18 and 19, catheter 1010 has a tip 1056 and bundle 
1095 operating and constructed as with the FIG. 15 embodiment except as 
follows. The bundle 1095 has a flat tip end 1095T from which beams 1038B 
pass from a plurality of fibers 1038 to a reflector 1096, which may be 
either a piece of silicon glass or a mirrored reflector. The reflector 
1096 may be made and operate in the fashion explained in Saadatmanesh et 
al. U.S. Pat. No. 5,242,438, issued Sep. 7, 1993 and hereby incorporated 
by reference except that the present reflector is shaped differently as 
discussed hereafter. 
The reflector 1096 has a series of parallel ridges 1096R and valleys 1096V 
extending across reflector 1096 perpendicular to the plane of view of 
FIGS. 18 and 19. The ridges and valleys are stepped in that those closest 
the one-hole window 1056W are further from tip end 1095T than the ridges 
and valleys which are further from the window 1056W. Since the slopes 
between ridges 1096R and valleys 1096V which face the window 1056W have a 
45 degree slope relative to beams 1038 (corresponding to the axes of 
fibers 1038), beams 1038 are reflected at right angles (90 degrees) 
becoming beams 1064. The beam widths for beams 1064 would be as discussed 
with respect to FIGS. 1-6. 
FIG. 20 shows a bottom view (i.e., side receiving laser energy) of an 
alternate reflector 1196 similar in operation and construction as 
reflector 1096 except as follows. Reflector 1196 has a plurality of rows 
1197 of width W, each row having stepped ridges 1196R and valleys 1196V 
such that a cross section along or parallel to any of lines 1197R (which 
separate adjacent rows) reveals a profile like that presented by reflector 
1096 in FIG. 19. By using a plurality of the rows 1197, a single optical 
fiber like fiber 38 of FIG. 2 (but without a prism at its tip) could apply 
a relatively wide beam to reflector 1196. The reflector 1196 would receive 
such a single beam (not shown) exiting in line with the central axis of 
the single optical fiber and create multiple beams by reflection. The 
reflection would be accomplished as discussed for FIG. 19 except that the 
reflection would actually be separating a relatively wide beam into a 
plurality of beams (not shown). The beam widths would be as discussed with 
respect to FIGS. 1-6. 
It will be readily appreciated that the embodiments of FIGS. 7 to 20 would 
be constructed in the same fashion as the FIGS. 1 to 6 embodiments except 
for the illustrated differences. All of the embodiments which heat the 
patient's cells in vivo without direct application of laser energy to the 
cells would heat the cells to 42 to 45 degrees centigrade for improving 
transfection receptivity. All of the embodiments may use one or two 
occluder balloons as discussed in connection with the FIG. 1 embodiment 
and may use feedback control as discussed with respect to FIG. 8. Although 
the discussion of FIGS. 6 to 20 has concentrated on DNA plasmids for 
treatment, drugs or other materials could be used. The treatment agent may 
have plasmids coprecipitated with CaPO.sub.4 or some other catalyst as 
that may be at least advisable to improve transfection. 
Tests by the present inventor of in vitro transfection into bovine aorta 
smooth muscle cells (SMC) have shown that heat can increase the efficiency 
of transfection. The plasmid pXGH5, which encodes a human growth hormone 
(hGH) reporter, was used to determine efficiency of transfection. SMC 
transfection was performed using DNA coprecipitated with CaPO.sub.4. SMCs 
were immediately heated for up to 1 hour at 42 to 45 degrees centigrade. 
Results were compared to paired control experiments conducted on the same 
day in the absence of heating. Transient gene expression was determined by 
radioimmunoassay of spent medium for hGH 48 hours after transfection. SMCs 
heated and exposed to DNA without CaPO.sub.4 showed no gene expression. In 
the 60 paired experiments, SMCs transfected by CaPO.sub.4 coprecipitation 
of DNA and heated demonstrated an increased hGH production compared to 
unheated controls, 20.3 plus or minus 3.7 versus 16.8 plus or minus 3.4 
(mean plus or minus SE) respectively, p&lt;0.002. 
With respect to the various embodiments applying laser energy to the 
tissues of a patient, one may use an external chromophore which will 
absorb wavelengths selectively to raise the temperature of the desired 
tissue. This would selectively heat the target tissue and reduce heat 
applied to other tissues. Also, it should smooth out the process of 
heating (i.e., avoid too fast temperature changes) and provide a more even 
temperature distribution over the desired tissues. Such a chromophore may 
involve oral or intravenous injection of materials which will concentrate 
in the desired tissues and make them more receptive to laser heating. 
Also, such materials to increase receptivity to laser heating may be 
inserted in the channel or channels of the catheter to be directly applied 
adjacent the heating. 
Although specific constructions and examples have been presented herein, it 
is to be understood that these are for illustrative purposes only. Various 
modifications and adaptations will be apparent to those of skill in the 
art. For example, one might use a double balloon catheter with the laser 
beams and treatment agents being applied from a window portion in between 
two balloons. Although the present invention has the highly advantageous 
feature of injecting the treatment agents out the same holes as the laser 
beams, the present invention, in its broadest aspects, might include a 
double balloon arrangement wherein the treatment agent comes out holes 
separate from the laser beams and fills the chamber established between 
the two balloons blocking part of an artery. Since the laser beams would 
be porating the cells within artery walls between two such balloons, the 
treatment agent, such as genetic material, could transfect the cells more 
quickly than in the prior art double balloon technique discussed in the 
background portion of this application since that prior art technique did 
not provide for cell porations. Such a double balloon technique may also 
use known dye materials inserted to enhance absorption of laser energy. 
Such materials are disclosed in the present inventors' prior U.S. Pat. 
Nos. 4,860,743 and 5,041,109 issued respectively on Aug. 29, 1989 and Aug. 
20, 1991 and hereby incorporated by reference. Although the laser 40 would 
preferably be a pulsed type laser which could use feedback control as 
discussed, one would especially want to use a thermocouple (for feedback 
control as discussed) to guard against overheating if the laser 40 was a 
continuous wave laser. In view of these and other possible modifications, 
it will be appreciated that the scope of the present invention should be 
determined by reference to the claims appended hereto.